Amino transferase and application thereof in preparation of chiral cyclic amide compound

Abstract

The invention provides an aminotransferase and an application of the aminotransferase in preparation of a chiral cyclic amide compound. The invention specifically provides an aminotransferase mutant, and compared with an amino acid sequence as shown in SEQ ID NO: 1, the aminotransferase mutant contains the difference of amino acid residues at one or more sites. The aminotransferase provided by the invention is high in catalytic yield, good in selectivity and good in application prospect.

Description

An aminotransferase and its application in the preparation of chiral cyclic amide compounds Technical Field

[0001] This invention belongs to the field of enzyme engineering technology, specifically relating to an aminotransferase and its application in the preparation of chiral cyclic amide compounds. Background Technology

[0002] Chiral cyclic amides are chiral precursors for a variety of drugs in clinical stages. Chemically, they require at least six reaction steps, which can lead to low yields, poor selectivity, or high costs. Enzymatic catalysis can shorten the process to two steps, while also offering high atom economy and being environmentally friendly.

[0003] Existing patents (CN114934082B and CN114686451B) involve the generation of 4-aminovaleric acid via amine dehydrogenase, followed by the intramolecular cyclization of the intermediate optical 4-aminovaleric acid to synthesize (S)-5-methyl-2-pyrrolidone. This process requires two steps, is complex, and hinders industrial application. Existing technologies disclose a natural aminotransferase derived from Bacillus megaterium. This aminotransferase catalyzes the production of the β-(S) chiral lactam isomer with a very low excess value (ee ~ 1%). Existing technology (CN116555366A) discloses a mutant with a selectivity of ee ≥ 80%, and existing technology (CN116555211A) discloses a mutant with ee ≥ 90% and a yield > 55%. The selectivity and yield of the enzyme catalysis in these existing technologies need further improvement. Developing mutants with high stereoselectivity, good catalytic activity, and better heat tolerance has high industrial application value. Summary of the Invention

[0004] The technical problem to be solved by this invention is to overcome the shortcomings of low yield and poor selectivity in the enzyme-catalyzed synthesis of cyclic amide compounds in existing technologies. Therefore, this invention provides an aminotransferase and its application in the preparation of chiral cyclic amide compounds. The enzyme-catalyzed synthesis of cyclic amide compounds provided in this application has good yield, good selectivity, high yield, good heat tolerance, and promising application prospects.

[0005] To solve the above-mentioned technical problems, the present invention provides the following technical solution:

[0006] This invention provides an aminotransferase mutant that, compared to the amino acid sequence shown in SEQ ID NO:1, contains amino acid residues differing at one or more of the following sites:

[0007] 7th, 8th, 9th, 11th, 12th, 15th, 16th, 18th, 19th, 20th, 21st, 22nd, 24th, 25th, 28th, 29th, 32nd, 34th, 37th, 40th, 48th, 51st, 56th, 58th, 59th, 6th 0th digit, 61st digit, 67th digit, 69th digit, 70th digit, 73rd digit, 76th digit, 77th digit, 79th digit, 80th digit, 81st digit, 84th digit, 86th digit, 87th digit, 88th digit, 89th digit, 90th digit, 96th digit, 99th digit, 103rd digit, 104th digit, 108th digit, 110th digit, 114th digit, 1st digit 19th, 120th, 121st, 122nd, 123rd, 124th, 125th, 134th, 135th, 137th, 148th, 149th, 150th, 151st, 152nd, 155th, 156th, 158th, 159th, 161st, 162nd 163rd, 164th, 165th, 168th, 170th, 171st, 172nd, 174th, 175th, 176th, 177th, 178th, 179th, 181st, 182nd, 185th, 187th, 191st, 197th, 198th, 199th 200th, 201st, 202nd, 204th, 205th, 207th, 209th, 210th, 217th, 218th, 221st, 224th, 225th, 226th, 229th, 237th, 240th, 241st, 242nd, 243rd, 24th 4th, 245th, 249th, 250th, 257th, 260th, 261st, 269th, 271st, 272nd, 273rd, 280th, 286th, 288th, 290th, 295th, 297th, 307th, 314th, 315th, 321st, 3rd 22nd, 323rd, 324th, 325th, 326th, 327th, 328th, 329th, 330th, 331st, 332nd, 337th, 346th, 349th, 350th, 351st, 352nd, 355th, 356th, 358th, 359th, ... 362nd, 365th, 369th, 370th, 372nd, 373rd, 374th, 376th, 380th, 387th, 396th, 397th, 398th, 399th, 400th, 401st, 403rd, 404th, 408th, 415th, 416thPositions 420, 423, 424, 427, 428, 429, 436, 437, 442, 448, 451, 456, 459, 463, 466, 467, 469, 472, and 474; and the aminotransferase mutant exhibits activity superior to that of the aminotransferase with the amino acid sequence shown in SEQ ID NO:1.

[0008] The difference may be the deletion, addition, or substitution of amino acids; substitution is preferred.

[0009] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following 7th position: A, C, D, F, G, H, I, L, M, N, P, Q, S, T, V, W, or Y.

[0010] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following 8th position: C, D, E, F, H, K, L, M, N, Q, R, T, V, W, or Y.

[0011] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following 9th position: A, C, D, E, G, H, I, K, L, R, S, T, V, W, or Y.

[0012] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following 11th position: A, C, D, F, G, H, I, K, L, M, N, P, Q, R, S, V, or Y.

[0013] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following 12th position: A, C, D, E, F, G, H, I, L, M, P, S, T, V, W, or Y.

[0014] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following 15th position: C, D, F, H, I, K, L, M, N, P, Q, R, S, W, or Y.

[0015] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following 16th position: A, E, F, G, I, K, L, M, N, Q, R, or S.

[0016] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following 18th position: C, D, E, G, I, M, N, P, Q, S, T, or V.

[0017] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following 19th position: A, C, D, E, F, G, H, M, N, P, S, T, W, or Y.

[0018] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following 20th position: A, C, D, G, I, K, L, M, N, P, Q, R, S, T, or V.

[0019] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following 21st position: A, C, D, F, G, H, I, K, M, P, Q, R, T, V, W, or Y.

[0020] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 22: A, C, D, E, F, G, H, I, K, L, N, Q, R, S, V, W, or Y.

[0021] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following 24th position: A, C, D, E, F, G, H, K, L, N, Q, R, W, or Y.

[0022] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 25: A, C, D, E, F, G, H, K, P, Q, R, S, T, V, or Y.

[0023] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following 28th position: C, D, E, F, H, I, K, L, M, P, R, S, T, or Y.

[0024] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following 29th position: C, D, F, L, P, Q, R, S, or Y.

[0025] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following 32nd position: A, C, D, E, F, G, H, K, L, M, P, R, S, T, V, W, or Y.

[0026] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following 34th position: C, D, E, F, G, H, L, M, Q, R, S, T, W, or Y.

[0027] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following 37th position: F, G, H, I, K, L, P, Q, R, S, V, or W.

[0028] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following 40th position: A, C, G, H, I, K, L, N, P, R, S, T, W, or Y.

[0029] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 48: C, D, E, F, G, M, N, Q, R, S, T, V, W, or Y.

[0030] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following 51st position: C, D, G, H, I, K, L, M, Q, R, T, V, W, or Y.

[0031] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 56: A, C, D, F, G, I, L, N, P, R, S, V, or Y.

[0032] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 58: D, E, F, G, H, I, K, N, R, S, T, V, W, or Y.

[0033] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 59: C, D, E, F, G, I, K, N, P, Q, R, S, T, V, W, or Y.

[0034] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following 60th position: A, C, D, G, H, I, K, F, M, N, P, Q, R, S, T, or W.

[0035] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following 61st position: A, D, E, F, G, H, I, K, L, M, P, Q, R, V, or W.

[0036] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 67: A, C, D, E, G, H, I, L, M, N, P, Q, R, S, T, W, or Y.

[0037] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following 69th position: A, D, F, H, I, K, M, N, P, R, S, T, W, or Y.

[0038] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following 70th position: A, C, D, E, F, H, I, L, M, P, Q, R, S, T, or W.

[0039] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following 73rd position: C, E, F, G, H, I, K, L, M, N, Q, R, S, V, W, or Y.

[0040] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following 76th position: A, C, D, E, F, G, H, I, L, N, P, Q, R, S, T, V, or W.

[0041] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following 77th position: A, C, D, F, G, H, K, L, M, N, P, Q, R, S, or Y.

[0042] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following 79th position: C, D, E, G, I, K, N, P, Q, R, S, T, V, W, or Y.

[0043] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following 80th position: A, C, E, F, G, H, I, K, L, M, N, P, R, T, V, W, or Y.

[0044] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following 81st position: C, E, I, K, L, N, P, Q, S, T, V, W, or Y.

[0045] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following 84th position: A, C, D, E, H, I, K, L, N, P, R, T, V, W, or Y.

[0046] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following 86th position: A, C, D, E, F, G, I, K, L, M, N, P, Q, R, S, T, V, or Y.

[0047] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following 87th position: A, C, E, F, G, H, K, L, M, N, P, Q, R, S, T, V, W, or Y.

[0048] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following 88th position: A, C, D, E, F, G, H, I, K, L, M, N, P, Q, R, V, W, or Y.

[0049] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following 89th position: A, D, E, F, G, H, I, L, P, S, T, V, or W.

[0050] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following 90th position: C, E, F, G, H, K, L, P, Q, S, T, V, W, or Y.

[0051] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 96: A, C, D, E, G, H, I, L, N, P, Q, R, S, T, V, W, or Y.

[0052] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following 99th position: A, D, F, G, H, I, N, P, Q, R, S, T, V, W, or Y.

[0053] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 103: A, D, F, G, H, I, K, L, P, Q, R, S, T, V, or Y.

[0054] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 104: A, C, E, F, H, I, L, M, N, Q, R, V, or W.

[0055] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 108: A, C, E, G, H, I, K, M, P, R, S, T, V, or W.

[0056] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 110: A, C, F, G, I, K, L, M, P, Q, R, T, V, W, or Y.

[0057] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 114: A, C, D, E, F, G, I, L, M, N, P, Q, R, S, V, W, or Y.

[0058] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 119: A, C, D, E, F, G, H, L, M, N, Q, R, V, W, or Y.

[0059] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 120: A, C, D, E, F, G, H, I, K, L, M, N, Q, R, S, W, or Y.

[0060] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 121: D, E, F, H, I, M, N, P, R, S, V, or Y.

[0061] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has position 122 as: A, D, F, G, H, I, K, L, M, P, T, V, W, or Y.

[0062] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has position 123 as: A, D, F, G, H, I, K, L, M, P, Q, R, S, V, or W.

[0063] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 124: C, D, E, G, I, K, L, N, P, Q, S, W, or Y.

[0064] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 125: A, C, D, F, G, H, I, L, M, N, Q, S, T, W, or Y.

[0065] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 134: A, D, E, F, H, I, K, M, P, Q, S, T, or W.

[0066] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 135: A, C, D, E, F, H, I, L, N, P, Q, R, S, T, V, or W.

[0067] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 137: A, D, F, G, H, I, K, L, M, P, R, T, or W.

[0068] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 148: C, D, E, F, G, H, I, K, L, M, N, P, S, T, V, W, or Y.

[0069] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 149: A, C, D, E, F, G, I, K, L, M, P, R, S, T, V, or W.

[0070] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 150: A, C, D, F, H, I, K, L, M, N, Q, R, S, T, W, or Y.

[0071] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 151: A, C, D, G, H, K, L, M, N, Q, R, S, T, V, or Y.

[0072] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 152: A, C, D, E, F, G, H, K, L, M, N, P, Q, R, S, V, W, or Y.

[0073] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 155: C, E, F, I, L, M, N, Q, R, V, or Y.

[0074] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 156: C, D, E, F, G, H, I, K, L, M, N, P, Q, R, S, T, V, or W.

[0075] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 158: A, C, D, E, F, G, H, I, K, L, M, N, P, R, S, T, W, or Y.

[0076] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 159: C, D, F, G, H, K, L, N, Q, R, S, V, W, or Y.

[0077] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 161: A, D, E, G, I, K, M, P, Q, R, S, T, V, W, or Y.

[0078] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 162: A, C, D, E, F, G, H, I, K, L, N, P, Q, S, T, V, W, or Y.

[0079] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 163: A, C, D, G, H, I, L, M, N, P, R, T, W, or Y.

[0080] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 164: A, C, D, E, F, G, H, I, K, L, M, N, P, Q, R, S, T, V, or W.

[0081] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 165: A, C, F, G, H, I, K, L, M, P, Q, R, V, W, or Y.

[0082] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 168: A, C, D, E, F, G, H, I, K, N, P, Q, R, S, W, or Y.

[0083] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 170: A, C, D, E, I, K, L, M, N, P, Q, R, S, V, W, or Y.

[0084] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 171: A, D, E, F, H, I, K, L, M, N, P, R, S, T, V, or Y.

[0085] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 172: A, C, D, E, F, H, I, K, M, P, Q, R, S, T, V, W, or Y.

[0086] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 174: A, D, F, G, H, I, L, M, N, P, Q, R, S, V, W, or Y.

[0087] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 175: A, C, D, E, F, G, H, I, K, L, M, N, P, Q, R, T, V, W, or Y.

[0088] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 176: A, C, D, E, G, H, I, K, L, M, N, P, Q, T, V, W, or Y.

[0089] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 177: A, C, D, E, F, G, H, I, K, L, M, N, P, Q, R, T, V, or W.

[0090] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 178: C, D, F, G, H, I, K, L, N, P, Q, R, S, T, V, W, or Y.

[0091] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 179: C, D, F, G, H, I, L, N, Q, R, S, T, V, W, or Y.

[0092] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 181: D, E, F, G, H, I, K, M, Q, S, or V.

[0093] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 182: A, C, E, F, H, I, L, P, Q, S, T, V, or Y.

[0094] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 185: A, C, D, E, F, H, K, L, M, N, P, Q, R, S, T, V, or W.

[0095] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 187: A, C, E, F, G, H, L, M, N, P, Q, R, T, or Y.

[0096] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 191: A, C, D, E, F, G, H, I, K, N, P, Q, S, T, V, or W.

[0097] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 197: A, D, E, F, G, H, I, K, L, N, P, Q, R, S, T, V, W, or Y.

[0098] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 198: A, C, E, G, H, I, K, M, P, Q, T, V, W, or Y.

[0099] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 199: A, C, D, E, F, G, H, I, K, L, M, N, P, R, S, T, W, or Y.

[0100] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 200: A, C, E, F, H, K, L, M, N, Q, R, T, V, W, or Y.

[0101] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 201: A, C, D, E, F, G, H, I, K, L, M, N, P, Q, R, T, V, W, or Y.

[0102] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 202: A, C, E, F, G, H, K, L, M, P, R, S, T, W, or Y.

[0103] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 204: A, C, D, F, G, H, I, K, L, Q, R, T, W, or Y.

[0104] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 205: A, C, D, E, F, G, H, I, K, M, N, P, Q, R, S, T, W, or Y.

[0105] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has position 207 as: A, E, G, I, M, N, P, Q, R, S, V, W, or Y.

[0106] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 209: A, D, F, G, I, K, N, Q, R, T, V, W, or Y.

[0107] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 210: A, D, E, F, G, H, I, K, M, P, R, T, or V.

[0108] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 217: A, C, D, E, F, G, H, I, M, N, P, Q, R, S, T, V, W, or Y.

[0109] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 218: A, D, E, F, G, H, K, L, M, N, P, S, or T.

[0110] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 221: A, C, D, E, F, H, I, L, M, N, P, Q, S, T, V, W, or Y.

[0111] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 224: A, C, D, F, G, H, K, L, P, Q, R, S, T, or V.

[0112] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 225: A, C, D, E, F, G, H, K, L, M, P, Q, S, T, V, W, or Y.

[0113] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 226: A, C, D, F, H, I, K, L, M, P, Q, R, S, T, or V.

[0114] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 229: A, C, D, F, G, H, I, K, N, P, Q, R, S, T, V, W, or Y.

[0115] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 237: A, C, D, F, G, H, I, K, L, M, N, P, S, T, V, or W.

[0116] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 240: A, E, F, G, H, K, L, M, P, S, V, W, or Y.

[0117] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 241: A, D, H, I, K, P, R, S, T, V, or Y.

[0118] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 242: C, D, F, G, H, I, K, L, N, S, T, V, W, or Y.

[0119] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 243: A, C, D, F, H, I, K, N, P, Q, S, T, V, or W.

[0120] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 244: A, C, E, F, G, H, I, K, L, N, Q, V, W, or Y.

[0121] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 245: C, D, E, F, G, H, I, K, L, M, N, P, R, S, T, V, W, or Y.

[0122] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 249: A, C, D, E, G, H, I, K, L, M, N, P, Q, R, S, T, V, or W.

[0123] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 250: A, C, D, G, H, I, M, Q, R, S, or T.

[0124] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 257: A, D, E, F, H, L, M, N, P, R, S, T, V, W, or Y.

[0125] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 260: C, D, E, F, G, H, L, N, P, Q, R, S, T, or Y.

[0126] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 261: A, C, D, F, G, H, K, N, P, Q, R, S, T, W, or Y.

[0127] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 269: A, C, E, F, I, K, L, M, N, P, Q, S, T, V, W, or Y.

[0128] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 271: C, D, E, F, G, H, K, L, M, S, T, or Y.

[0129] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has position 272 as: A, C, E, G, P, Q, R, T, W, or Y.

[0130] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 273: D, E, F, G, H, I, K, L, M, N, P, Q, R, S, or Y.

[0131] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 280: C, F, G, H, I, L, M, N, P, Q, S, T, V, or W.

[0132] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 286: A, C, E, F, G, I, K, N, Q, R, S, T, V, W, or Y.

[0133] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 288: A, C, D, E, I, K, M, N, Q, R, S, V, W, or Y.

[0134] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 290: A, C, D, E, F, G, H, I, K, L, M, P, R, S, T, V, W, or Y.

[0135] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 295: A, C, D, E, F, G, H, I, K, L, M, N, P, Q, S, V, or Y.

[0136] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 297: A, C, D, E, K, L, M, N, Q, R, S, V, or W.

[0137] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 307: C, D, E, F, G, H, I, K, L, M, N, P, Q, S, T, V, or W.

[0138] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 314: A, D, E, F, G, H, I, L, M, N, P, Q, R, S, T, V, or W.

[0139] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 315: C, D, F, G, H, I, K, M, N, Q, R, S, T, V, W, or Y.

[0140] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 321: A, C, E, F, G, H, I, K, L, M, N, P, Q, R, S, T, V, or Y.

[0141] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has position 322 as: A, C, E, F, G, H, I, M, N, P, R, S, T, V, or W.

[0142] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has position 323 as: A, C, D, E, F, G, K, L, N, P, Q, R, V, W, or Y.

[0143] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 324: C, F, G, I, K, L, M, N, P, Q, S, or Y.

[0144] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 325: C, E, F, H, I, K, L, M, N, P, Q, S, T, V, or Y.

[0145] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 326: C, F, G, H, K, L, M, P, R, S, T, or V.

[0146] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 327: C, D, E, F, G, H, K, L, M, N, P, Q, R, T, V, W, or Y.

[0147] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 328: A, C, D, E, F, G, H, I, K, L, M, P, Q, R, S, T, W, or Y.

[0148] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has position 329 as: A, C, E, G, I, L, N, P, Q, R, T, V, W, or Y.

[0149] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 330: A, C, D, E, F, G, H, I, L, M, P, Q, S, V, or W.

[0150] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 331: A, C, D, E, F, G, H, I, K, L, M, N, P, Q, R, S, T, V, or W.

[0151] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 332: D, E, F, H, I, K, N, P, Q, R, S, T, W, or Y.

[0152] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 337: C, D, E, F, G, H, I, K, L, M, N, P, Q, R, S, T, V, W, or Y.

[0153] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 346: A, C, D, F, G, H, I, K, N, P, Q, T, V, or Y.

[0154] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 349: D, E, F, H, I, L, N, P, Q, T, V, W, or Y.

[0155] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 350: C, D, F, G, I, K, L, M, P, Q, R, S, T, V, W, or Y.

[0156] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 351: A, C, F, H, I, K, N, P, Q, R, S, T, or W.

[0157] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following sequence at position 352: C, D, E, F, H, I, K, L, P, Q, R, S, T, V, or W.

[0158] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has position 355 as: A, C, H, I, L, N, P, Q, R, T, W, or Y.

[0159] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 356: A, D, E, G, H, I, N, Q, S, T, V, W, or Y.

[0160] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 358: C, E, F, H, I, M, N, P, R, S, T, V, or W.

[0161] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 359: A, E, F, G, H, I, K, N, P, Q, R, S, T, V, W, or Y.

[0162] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 362: A, C, F, G, H, I, K, M, N, P, R, S, T, or Y.

[0163] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 365: A, C, D, E, F, G, H, I, L, N, P, Q, S, T, V, W, or Y.

[0164] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following sequence at position 369: D, F, K, L, M, N, Q, R, S, T, V, W, or Y.

[0165] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 370: C, D, E, F, G, I, K, M, N, P, Q, R, S, T, V, W, or Y.

[0166] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 372: A, C, D, E, F, G, I, K, L, M, N, P, R, S, T, V, W, or Y.

[0167] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has position 373 as: A, C, D, F, G, H, I, L, M, N, P, Q, V, or Y.

[0168] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 374: A, C, D, F, H, M, N, P, Q, R, T, V, W, or Y.

[0169] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following sequence at position 376: D, E, G, I, M, N, Q, R, T, W, or Y.

[0170] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 380: A, C, D, E, F, G, H, I, K, L, M, P, Q, S, T, V, or Y.

[0171] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 387: A, C, D, E, F, G, H, I, K, M, N, P, Q, R, S, T, V, W, or Y.

[0172] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 396: A, C, E, F, G, H, I, L, M, N, P, Q, S, T, W, or Y.

[0173] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 397: A, C, D, E, F, G, H, I, K, M, N, R, S, V, or W.

[0174] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 398: A, C, D, E, F, G, H, I, L, M, N, P, R, T, V, W, or Y.

[0175] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has position 399 as: A, C, E, F, H, L, M, N, P, Q, R, S, V, W, or Y.

[0176] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 400: A, C, D, E, F, G, H, I, L, M, N, R, S, W, or Y.

[0177] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 401: A, D, E, F, H, I, K, L, N, P, S, T, or W.

[0178] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 403: A, C, D, E, F, G, H, I, M, N, P, Q, R, S, T, V, W, or Y.

[0179] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has position 404 as: A, C, D, G, K, M, N, R, T, V, W, or Y.

[0180] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 408: A, C, D, E, G, H, I, K, L, M, N, P, Q, R, S, T, or V.

[0181] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has position 415 as: A, C, D, E, F, G, H, I, K, L, M, P, Q, S, V, or Y.

[0182] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 416: A, C, D, F, G, H, I, K, L, M, N, P, R, S, T, or W.

[0183] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has position 420 as: A, D, E, G, H, K, L, P, R, S, W, or Y.

[0184] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has position 423 as: A, C, F, G, I, K, L, M, N, Q, R, S, T, V, W, or Y.

[0185] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 424: D, E, F, G, L, M, N, P, Q, R, S, V, W, or Y.

[0186] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 427: A, C, D, E, F, G, H, I, K, M, N, P, R, S, T, V, W, or Y.

[0187] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 428: C, D, F, G, K, L, M, N, P, R, or T.

[0188] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 429: A, C, D, F, G, H, I, L, M, N, P, Q, R, S, T, V, W, or Y.

[0189] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 436: A, C, D, E, F, G, H, I, K, L, N, P, Q, R, S, T, W, or Y.

[0190] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has position 437 as: A, C, D, E, F, G, H, I, L, N, P, Q, R, S, T, or Y.

[0191] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has position 442 as: A, C, E, F, G, H, I, L, M, N, Q, S, T, V, W, or Y.

[0192] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 448: A, C, D, E, F, G, H, I, K, L, M, P, Q, R, S, T, or Y.

[0193] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 451: A, C, D, E, H, I, K, M, N, P, Q, S, T, V, W, or Y.

[0194] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 456: D, E, F, G, H, I, L, N, P, Q, R, S, T, V, or Y.

[0195] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 459: A, C, E, F, G, L, M, N, P, R, S, V, W, or Y.

[0196] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 463: A, C, D, E, F, G, H, I, L, M, N, P, Q, R, S, T, V, or W.

[0197] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 466: A, C, E, F, G, H, I, K, L, M, N, P, Q, R, T, V, or Y.

[0198] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 467: A, C, E, F, G, H, I, K, L, M, N, Q, R, S, T, or V.

[0199] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has position 469 as: A, C, D, F, H, I, K, L, M, N, P, S, T, V, W, or Y.

[0200] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 472: A, C, D, G, H, K, L, M, N, P, S, T, W, or Y.

[0201] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has the following amino acid sequence at position 474: A, C, D, E, F, G, H, I, K, N, P, R, S, T, V, W, or Y.

[0202] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, has any of the following combinations of amino acid residue differences:

[0203]

[0204]

[0205]

[0206]

[0207]

[0208]

[0209]

[0210]

[0211]

[0212]

[0213]

[0214]

[0215]

[0216]

[0217]

[0218] In this context, " / " represents s and .

[0219] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, contains differences in amino acid residues at the following sites: 007D and 073N, and position 060 is F or M. Preferably, the aminotransferase mutant further contains differences in amino acid residues at one or more of the following sites: 076R, 088I, 096R, 156R, 158T, 162K, 164M, and 170A. Any of the above aminotransferase mutants, compared to the amino acid sequence shown in SEQ ID NO:1, may further contain differences in amino acid residues at one or more of the following sites: 175E, 178Q, 197R, 199R, 201G, 205Y, 221D, 245T, 249R, 290H, 295I, and 327T.

[0220] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, contains amino acid residue differences at one or more of the following sites: 086W, 175E, 199T, 201G, 221D, 249R, 328I, 370D, 380S, 387M, 436T, 448N, with I or R at position 088, M or L at position 164, and W or F at position 331.

[0221] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, contains differences in amino acid residues at the following sites: 007D, 021I, 073N, 076R, 096R, 156R, 158T, 162K, 170A, 178Q, and 197R; preferably, the aminotransferase mutant further contains differences in amino acid residues at the following sites: 205Y, 245T, 290H, 295I, 327T, and 337C; more preferably, the aminotransferase mutant further contains differences in amino acid residues at the following sites: 372S, 403P, 408A, and 442I. Compared with the amino acid sequence shown in SEQ ID NO:1, any of the above amino acid transferase mutants may further contain differences in one or more of the following amino acid residues: S, M, or F at position 060; V, I, H, N, or R at position 088; R or T at position 199; A or D at position 211; K or R at position 249; M or I at position 328; G or S at position 380; and L or T at position 436.

[0222] In one embodiment, the aminotransferase mutant, compared to the amino acid sequence shown in SEQ ID NO:1, contains amino acid residue differences at the following sites: 007D, 021I, 060 as S, F, or M; 073N, 076R, 088 as V, I, N, H, or R; 096R, 156R, 158T, 162K, 170A, 178Q, 197R, 199 as R or T; 205Y, 221 as A or D; 245T, 249 as K or R. The amino acid differences at positions 290H, 295I, 327T, 328 (M or I), 337C, 372S, 380 (G or S), 403P, 408A, 436 (L or T), and 442I are preferred. The amino acid differences at one or more of the following sites are also preferred: 086T, 164 (M or L), 175E, 176W, 177A, 201G, 331 (F or W), 370 (D or E), 387M, and 448A.

[0223] Preferably, the aminotransferase mutant has any of the following combinations of amino acid residue differences compared to the amino acid sequence shown in SEQ ID NO:1:

[0224] Mutant 18: 007D / 021I / 060S / 073N / 076R / 086T / 088V / 096R / 156R / 158T / 162K / 164M / 170A / 178Q / 197R / 199R / 205Y / 221A / 245T / 249K / 290H / 295I / 327T / 328M / 337C / 372S / 380G / 403P / 408A / 436L / 442I / 448A;

[0225] Mutant 49: 007D / 021I / 060S / 073N / 076R / 086T / 088V / 096R / 156R / 158T / 162K / 170A / 178Q / 197R / 199R / 205Y / 221A / 245T / 249K / 290H / 295I / 327T / 328M / 337C / 370E / 372S / 380G / 403P / 408A / 436L / 442I / 448A;

[0226] Mutant 73: 007D / 021I / 060S / 073N / 076R / 086T / 088V / 096R / 156R / 158T / 162K / 170A / 178Q / 197R / 199R / 205Y / 221A / 245T / 249K / 290H / 295I / 327T / 328M / 337C / 372S / 380G / 403P / 408A / 436L / 442I / 448A;

[0227] Mutant 102: 007D / 021I / 060F / 073N / 076R / 088I / 096R / 156R / 158T / 162K / 164M / 170A / 175E / 178Q / 197R / 199R / 201G / 205Y / 221D / 245T / 249R / 290H / 295I / 327T / 328I / 331F / 337C / 372S / 380S / 387M / 403P / 408A / 436T / 442I;

[0228] Mutant 231: 007D / 021I / 060M / 073N / 076R / 088I / 096R / 156R / 158T / 162K / 164M / 170A / 175E / 178Q / 197R / 199R / 201G / 205Y / 221D / 245T / 249R / 290H / 295I / 327T / 328M / 331W / 337C / 372S / 380S / 387M / 403P / 408A / 436T / 442I;

[0229] Mutant 245: 007D / 021I / 060S / 073N / 076R / 088V / 096R / 156R / 158T / 162K / 170A / 178Q / 197R / 199R / 205Y / 221A / 245T / 249K / 290H / 295I / 327T / 328M / 337C / 372S / 380G / 403P / 408A / 436L / 442I / 448A;

[0230] Mutant 256: 007D / 021I / 060S / 073N / 076R / 086T / 088V / 096R / 156R / 158T / 162K / 164L / 170A / 178Q / 197R / 199R / 205Y / 221A / 245T / 249K / 290H / 295I / 327T / 328M / 337C / 372S / 380G / 403P / 408A / 436L / 442I / 448A;

[0231] Mutant 280: 007D / 021I / 060S / 073N / 076R / 088I / 096R / 156R / 158T / 162K / 164M / 170A / 175E / 178Q / 197R / 199R / 201G / 205Y / 221D / 245T / 249R / 290H / 295I / 327T / 328I / 331W / 337C / 372S / 380S / 387M / 403P / 408A / 436T / 442I;

[0232] Mutant 289: 007D / 021I / 060S / 073N / 076R / 088I / 096R / 156R / 158T / 162K / 170A / 175E / 178Q / 197R / 199R / 201G / 205Y / 221D / 245T / 249R / 290H / 295I / 327T / 328M / 337C / 372S / 380S / 403P / 408A / 436T / 442I;

[0233] Mutant 301: 007D / 021I / 060S / 073N / 076R / 088I / 096R / 156R / 158T / 162K / 170A / 175E / 176W / 178Q / 197R / 199R / 205Y / 221D / 245T / 249R / 290H / 295I / 327T / 328M / 337C / 372S / 380S / 403P / 408A / 436L / 442I;

[0234] Mutant 338: 007D / 021I / 060S / 073N / 076R / 088N / 096R / 156R / 158T / 162K / 164M / 170A / 175E / 176W / 177A / 178Q / 197R / 199R / 201G / 205Y / 221D / 245T / 249R / 290H / 295I / 327T / 328M / 331W / 337C / 370D / 372S / 380S / 403P / 408A / 436T / 442I;

[0235] Mutant 344: 007D / 021I / 060F / 073N / 076R / 088I / 096R / 156R / 158T / 162K / 164M / 170A / 175E / 178Q / 197R / 199R / 201G / 205Y / 221D / 245T / 249R / 290H / 295I / 327T / 328M / 331W / 337C / 372S / 380S / 387M / 403P / 408A / 436T / 442I;

[0236] Mutant 356: 007D / 021I / 060S / 073N / 076R / 086T / 088V / 096R / 156R / 158T / 162K / 170A / 178Q / 197R / 199R / 205Y / 221A / 245T / 249K / 290H / 295I / 327T / 328M / 337C / 372S / 380G / 403P / 408A / 436L / 442I;

[0237] Mutant 368: 007D / 021I / 060S / 073N / 076R / 088I / 096R / 156R / 158T / 162K / 170A / 175E / 178Q / 197R / 199R / 205Y / 221D / 245T / 249R / 290H / 295I / 327T / 328M / 331W / 337C / 372S / 380S / 403P / 408A / 436L / 442I;

[0238] Mutant 371: 007D / 021I / 060S / 073N / 076R / 088I / 096R / 156R / 158T / 162K / 164M / 170A / 175E / 178Q / 197R / 199R / 201G / 205Y / 221D / 245T / 249R / 290H / 295I / 327T / 328M / 331W / 337C / 372S / 380S / 387M / 403P / 408A / 436T / 442I;

[0239] Mutant 398: 076D / 086I / 096T / 177G / 201A / 205Q / 295L / 337W / 370K / 380A / 387A;

[0240] Mutant 406: 007D / 021I / 060S / 073N / 076R / 088I / 096R / 156R / 158T / 162K / 170A / 175E / 178Q / 197R / 199R / 201G / 205Y / 221D / 245T / 249R / 290H / 295I / 327T / 328M / 337C / 372S / 380S / 403P / 408A / 436L / 442I;

[0241] Mutant 418: 007D / 021I / 060S / 073N / 076R / 088V / 096R / 156R / 158T / 162K / 170A / 175E / 178Q / 197R / 199R / 205Y / 221D / 245T / 249R / 290H / 295I / 327T / 328M / 337C / 372S / 380S / 403P / 408A / 436L / 442I;

[0242] Mutant 423: 007D / 021I / 060S / 073N / 076R / 088H / 096R / 156R / 158T / 162K / 170A / 175E / 178Q / 197R / 199R / 201G / 205Y / 221D / 245T / 249R / 290H / 295I / 327T / 328M / 337C / 372S / 380S / 403P / 408A / 436L / 442I;

[0243] Mutant 440: 007D / 021I / 060S / 073N / 076R / 088R / 096R / 156R / 158T / 162K / 164M / 170A / 175E / 178Q / 197R / 199T / 201G / 205Y / 221D / 245T / 249R / 290H / 295I / 327T / 328I / 331F / 337C / 372S / 380S / 387M / 403P / 408A / 436T / 442I;

[0244] Mutant 441: 007D / 021I / 060S / 073N / 076R / 088I / 096R / 156R / 158T / 162K / 170A / 175E / 178Q / 197R / 199R / 205Y / 221D / 245T / 249R / 290H / 295I / 327T / 328M / 337C / 372S / 380S / 403P / 408A / 436L / 442I.

[0245] This invention provides an enzyme preparation containing the aminotransferase mutant described in any embodiment of the invention. The enzyme preparation may be a buffer solution containing the aminotransferase mutant. Preferably, the buffer solution is sodium borate buffer, phosphate buffer (PBS), trimethylolpropionate buffer (TRIS), or triethanolamine buffer (TOEA).

[0246] In one or more embodiments, the enzyme preparation is a powder, such as a lyophilized powder.

[0247] This invention provides an isolated polynucleotide encoding an aminotransferase mutant as described in any embodiment of this invention. Preferably, it further comprises a complementary sequence to the polynucleotide sequence.

[0248] This invention provides a nucleic acid construct comprising the aforementioned polynucleotides; preferably, the nucleic acid construct is an expression cassette or vector, such as a cloning vector or expression vector. Preferably, the nucleic acid construct is a recombinant expression vector comprising the aforementioned polynucleotides.

[0249] This invention provides a transformant comprising the aforementioned polynucleotide or nucleic acid construct. Preferably, the host cell used in the construction of the transformant is *Escherichia coli*.

[0250] The present invention provides a method for preparing the aminotransferase mutant, the method comprising: culturing the above-mentioned transformant to obtain the aminotransferase mutant.

[0251] This invention also provides the application of the above-mentioned aminotransferase mutant or enzyme preparation in the preparation of chiral lactam compounds, wherein, in the presence of an amino donor, the aminotransferase mutant catalyzes the compound represented by formula I. Synthetic chiral lactam compounds;

[0252] R3 is an oxo group (=O) or -CHO;

[0253] R1 and R2 are independently C 1-6 Alkyl or C 6-14 Aryl.

[0254] The present invention also provides a method for preparing a lactam compound, comprising the following steps: in the presence of the aminotransferase mutant, subjecting a compound of Formula I and an amino donor to an amination reaction to obtain a lactam compound;

[0255]

[0256] R3 is an oxo group (=O), -CHO, or...

[0257] Y1 and Y2 are independently H or -SO2X, where X is H or an alkali metal;

[0258] R1 and R2 are independently C 1-6 Alkyl, C 6-14 aryl or C substituted with one or more R4 6-14 Aryl;

[0259] R4 is independently C 1-6 Alkyl, C 1-6 Alkyl, halogen, or hydroxyl;

[0260] When R3 is an oxo group or When the lactam compound is

[0261] When R3 is -CHO, the lactam compound is And the asterisk (*) marks C as S configuration.

[0262] In one scheme, R3 is an oxo group (=O) or -CHO.

[0263] In one particular scheme, R2 is C 1-6 alkyl.

[0264] In one scheme, R3 is C 1-6 Alkyl or aryl.

[0265] In one scheme, the C 1-6 The alkyl group is methyl, ethyl, isopropyl, n-propyl, n-butyl, tert-butyl, sec-butyl, isobutyl, or tert-butyl, for example, methyl, ethyl, isopropyl, n-propyl, or tert-butyl.

[0266] In one embodiment, Y1 is H, Y2 is -SO2X, and X is H or an alkali metal. The alkali metal can be Na, K, or Li, for example, Na.

[0267] In one of the schemes, for

[0268] In one embodiment, the aryl group is C 6-14 Aryl, preferably phenyl or naphthyl, such as phenyl.

[0269] In one embodiment, the compound shown in Formula I is

[0270] In one of the solutions, for

[0271] In one of the solutions, for

[0272] The amino donor is a conventional amino donor in the art. Preferably, the amino donor is an amine compound or its salt, such as one or more of isopropylamine, L-alanine, DL-alanine, L-lysine, DL-lysine, (S)-1-phenylethylamine, (R)-1-phenylethylamine, (RS)-1-phenylethylamine, pentanediamine, o-phenylenediamine, o-phenylenediamine hydrochloride, 2-(4-nitrobenzene)ethylamine, and 2-(4-nitrobenzene)ethylamine hydrochloride; preferably, the amino donor is isopropylamine.

[0273] The molar ratio of the amino donor to the compound shown in Formula I is conventional in the art; preferably (1 to 6):1, for example 4:1.

[0274] In one embodiment, the reaction solvent for the amination reaction is a buffer solution, or a mixture of a buffer solution and a conventional organic solvent in the art, wherein the organic solvent is methyl tert-butyl ether, isopropyl ether, tetrahydrofuran, 2-methyltetrahydrofuran, 1,4-dioxane, ethyl acetate, dimethyl sulfoxide, N,N-dimethylformamide, methanol, ethanol, isopropanol, or n-butanol; preferably, the reaction solvent for the amination reaction is a buffer solution.

[0275] In one embodiment, when the reaction solvent for the amination reaction is a mixture of a buffer solution and a conventional organic solvent in the art, the organic solvent accounts for 0 to 250 mL / L of the total reaction volume.

[0276] The buffer solution is a conventional buffer solution in the art, preferably one or more of sodium borate buffer, phosphate buffer (PBS), trimethylolpropionate buffer (TRIS), and triethanolamine buffer (TEOA); more preferably sodium borate buffer, trimethylolpropionate buffer, or triethanolamine buffer.

[0277] The volume-to-mass ratio of the buffer solution to the compound of Formula I is a conventional volume-to-mass ratio in the art; preferably 5 to 150 mL / g, for example 20 to 110 mL / g, more preferably 28 mL / g, 50 mL / g or 100 mL / g.

[0278] The pH value of the amination reaction is a conventional pH value in the art, preferably 9.0 to 11; more preferably 9.0 to 10.5; for example 9.0, 9.5, 10.0, 10.5 or 11.0.

[0279] In one embodiment, the reaction temperature of the amination reaction is 25°C to 50°C; more preferably, it is 47°C, 37°C, 55°C or 27°C.

[0280] The amount of the aminotransferase mutant used is the conventional amount used in such reactions in the art, for example, the mass ratio of the aminotransferase mutant to the compound shown in Formula I is (0.01-1):1, such as 0.05:1 or 0.1:1.

[0281] The reaction time of the amination reaction is a conventional reaction time in the art. The reaction time is usually related to the reaction scale and the reaction process can be monitored by conventional means in the art. Generally, the reaction endpoint is when the compound shown in Formula I reacts completely or stops reacting. The reaction time is preferably 16 to 20 hours; for example, 16 hours, 18 hours or 20 hours.

[0282] In one embodiment, the amination reaction includes the following steps: adding the compound of Formula I and the aminotransferase mutant to a mixture containing the buffer solution and the amino donor, mixing thoroughly, and then reacting at 25°C to 50°C.

[0283] In one embodiment, the amination reaction is followed by the following post-processing steps: after the reaction is complete, the pH is adjusted (e.g., by adding alkali to adjust the pH to 13), water and ether solvents are added for extraction and separation, the organic phase is dried (e.g., by drying with anhydrous sodium sulfate), and the ether solvents are removed (e.g., by vacuum distillation).

[0284] In one embodiment, the ether solvent is a conventional ether solvent in the art, preferably tert-butyl methyl ether.

[0285] In one embodiment, the drying process employs conventional drying methods in the art, preferably drying with a desiccant; the desiccant is a conventional desiccant in the art, preferably anhydrous sodium sulfate.

[0286] In one embodiment, the alkali is a conventional alkali in the art, preferably sodium hydroxide or potassium hydroxide, such as potassium hydroxide.

[0287] In one embodiment, the method for removing ether solvents is a conventional method in the art, preferably vacuum distillation, for example, vacuum distillation at a pressure of -0.09 MPa.

[0288] Without violating common sense in the field, the above-mentioned preferred conditions can be combined arbitrarily to obtain various preferred embodiments of the present invention.

[0289] The reagents and raw materials used in this invention are all commercially available.

[0290] Unless otherwise specified, the abbreviations of amino acids used in this invention are conventional in the art, and the specific abbreviations corresponding to the amino acids are shown in the table below:

[0291]

[0292]

[0293] The positive and progressive effects of this invention are as follows:

[0294] This invention utilizes genetic engineering to direct the evolution of an aminotransferase, resulting in several mutants with higher activity, stereospecificity, and thermal stability. Based on these aminotransferase mutants, this invention develops a one-step synthesis process for chiral lactams from isopropyl levulinate compounds. This process exhibits good heat resistance (e.g., capable of reacting at relatively high temperatures (47°C)), high yield (e.g., greater than 80%), high purity (e.g., greater than 98.0%), and good selectivity. Detailed Implementation

[0295] The present invention is further illustrated below by way of embodiments, but the invention is not limited to the scope of the embodiments described herein. Experimental methods in the following embodiments that do not specify specific conditions were performed according to conventional methods and conditions, or as selected according to the product instructions.

[0296] Preparation Example: Preparation of Enzyme Mutants

[0297] Mutations were performed on the wild-type aminotransferase (nucleotide sequence SEQ ID NO:2) as shown in Table 1 below to obtain the enzyme mutants of the present invention.

[0298] Table 1

[0299]

[0300]

[0301]

[0302]

[0303]

[0304]

[0305]

[0306]

[0307]

[0308]

[0309]

[0310]

[0311]

[0312]

[0313]

[0314] The configuration of the target product in the embodiments was confirmed based on the S-configuration standard. Under the same conditions, the retention time of the peak of the target product in the embodiments was consistent with the retention time of the peak of the S-configuration standard.

[0315] The analytical methods and results of the target product in Example 1 are shown in the table below.

[0316]

[0317]

[0318] In this embodiment, the analytical method is the same as that in Example 1, and the retention times of each compound are shown in the table below:

[0319]

[0320] Example 1

[0321] In sodium borate buffer, aminotransferase catalyzes the synthesis of lactam from compound 2.

[0322]

[0323] Add 0.06 g of pyridoxal phosphate and 90 mL of 0.05 mol / L sodium borate buffer (pH 11) containing isopropylamine to a 100 mL beaker, and stir until the pyridoxal phosphate is completely dissolved to obtain a homogeneous solution. Transfer 100.0 mL of the solution from the beaker to a 250 mL reaction flask, and add 1 g of the compound and 0.1 g of aminotransferase containing mutant 289 to the solution. Vortex to mix the resulting solution and react at 47 °C for 20 h. After the reaction was completed, the reaction solution was removed, and 10 mL of water was added. The solution was then extracted with 200 mL of methyl tert-butyl ether (pH adjusted to 13 with potassium hydroxide). After extraction, the organic phase was removed and anhydrous sodium sulfate was added to remove water. The solution was then filtered to obtain the filtrate, which was then distilled under reduced pressure at -0.09 MPa (gauge pressure) and purified by column chromatography. The final product was 0.642 g, with a yield of 97.7%. The product was analyzed by NMR, and the results showed that the purity of the chiral lactam was greater than 98%.

[0324] Example 2

[0325] In sodium borate buffer, aminotransferase catalyzes the synthesis of lactam from compound 3.

[0326]

[0327] 0.06 g of pyridoxal phosphate and 90 mL of 0.05 mol / L sodium borate buffer (pH 11) containing isopropylamine were added to a 100 mL beaker. The mixture was stirred until pyridoxal phosphate was completely dissolved to obtain a homogeneous solution. 50 mL of the solution was transferred from the beaker to a 200 mL reaction flask, and 1 g of the compound and 0.1 g of aminotransferase with mutant 371 were added. The mixture was vortexed to mix the solution and then reacted at 47 °C for 20 h. After the reaction, the reaction solution was removed, and 10 mL of water was added. The mixture was then extracted with 200 mL of methyl tert-butyl ether (pH adjusted to 13 with potassium hydroxide). After extraction, the organic phase was removed, and anhydrous sodium sulfate was added to remove water. The filtrate was then filtered and distilled under reduced pressure at -0.09 MPa (gauge pressure). The filtrate was purified by column chromatography to obtain 0.650 g of the product, with a yield of 99%. NMR analysis showed that the purity of the chiral lactam was greater than 98%.

[0328] Example 3

[0329] In sodium borate buffer, aminotransferase catalyzes compound 4.

[0330]

[0331] 0.06 g of pyridoxal phosphate and 90 mL of 0.05 mol / L sodium borate buffer (pH 11) containing isopropylamine were added to a 100 mL beaker and stirred until the pyridoxal phosphate was completely dissolved to obtain a homogeneous solution. 100 mL of the solution was transferred from the beaker to a 250 mL reaction flask, and 1 g of the compound and 0.1 g of aminotransferase with mutant 338 were added to the solution. The mixture was vortexed and incubated at 37 °C for 20 h. After the reaction, the solution was removed, 10 mL of water was added, and the mixture was extracted with 200 mL of methyl tert-butyl ether (pH adjusted to 13 with potassium hydroxide). After extraction, the organic phase was removed, and anhydrous sodium sulfate was added to remove water. The filtrate was then filtered and distilled under reduced pressure at -0.09 MPa (gauge pressure). The product was purified by column chromatography, yielding 0.624 g of the product (95% yield). NMR analysis showed that the purity of the chiral lactam was greater than 98%.

[0332] Example 4

[0333] In sodium borate buffer, aminotransferase catalyzes the synthesis of lactam from compound 1.

[0334]

[0335] 0.06 g of pyridoxal phosphate and 90 mL of 0.05 mol / L sodium borate buffer (pH 11) containing isopropylamine were added to a 100 mL beaker. The mixture was stirred until pyridoxal phosphate was completely dissolved to obtain a homogeneous solution. 50 mL of the solution was transferred from the beaker to a 200 mL reaction flask, and 1 g of the compound and 0.1 g of aminotransferase with mutant 102 were added. The mixture was vortexed to mix the solution and then reacted at 47 °C for 20 h. After the reaction, the reaction solution was removed, and 10 mL of water was added. The mixture was then extracted with 200 mL of methyl tert-butyl ether (pH adjusted to 13 with potassium hydroxide). After extraction, the organic phase was removed, and anhydrous sodium sulfate was added to remove water. The filtrate was then filtered and distilled under reduced pressure at -0.09 MPa (gauge pressure). The filtrate was purified by column chromatography to obtain 0.597 g of the product, with a yield of 95.4%. NMR analysis showed that the purity of the chiral lactam was greater than 98%.

[0336] Example 5

[0337] In sodium borate buffer, aminotransferase catalyzes the synthesis of lactam from compound 1.

[0338]

[0339] 0.06 g of pyridoxal phosphate and 90 mL of 0.05 mol / L sodium borate buffer (pH 10) containing isopropylamine were added to a 100 mL beaker and stirred until the pyridoxal phosphate was completely dissolved to obtain a homogeneous solution. 100 mL of the solution was transferred from the beaker to a 250 mL reaction flask, and 1 g of the compound and 0.1 g of aminotransferase with mutant 440 were added to the solution. The mixture was vortexed and incubated at 47 °C for 20 h. After the reaction, the solution was removed, and 10 mL of water was added. Extraction was performed with 200 mL of methyl tert-butyl ether (pH adjusted to 13 with potassium hydroxide). After extraction, the organic phase was removed, and anhydrous sodium sulfate was added to remove water. The filtrate was then filtered and distilled under reduced pressure at -0.09 MPa (gauge pressure). The product was purified by column chromatography, yielding 0.617 g of the product (98.5%). NMR analysis showed that the purity of the chiral lactam was greater than 98%.

[0340] Example 6

[0341] In sodium borate buffer, aminotransferase catalyzes the synthesis of lactam from compound 4.

[0342]

[0343] 0.06 g of pyridoxal phosphate and 90 mL of 0.05 mol / L sodium borate buffer (pH 10) containing isopropylamine were added to a 100 mL beaker. The mixture was stirred until all pyridoxal phosphate was dissolved to obtain a homogeneous solution. 50 mL of the solution was transferred from the beaker to a 200 mL reaction flask, and 1 g of the compound and 0.1 g of aminotransferase containing mutant 301 were added. The mixture was vortexed to mix the solution and then reacted at 47 °C for 20 h. After the reaction, the reaction solution was removed, and 10 mL of water was added. The mixture was then extracted with 200 mL of methyl tert-butyl ether (pH adjusted to 13 with potassium hydroxide). After extraction, the organic phase was removed, and anhydrous sodium sulfate was added to remove water. The filtrate was then filtered and distilled under reduced pressure at -0.09 MPa (gauge pressure). The product was purified by column chromatography, yielding 0.64 g of the product (97.4%). NMR analysis showed that the purity of the chiral lactam was greater than 98%.

[0344] Example 7

[0345] In sodium borate buffer, aminotransferase catalyzes the synthesis of lactam from compound 5.

[0346]

[0347] 0.06 g of pyridoxal phosphate and 90 mL of 0.05 mol / L sodium borate buffer (pH 10.5) containing isopropylamine were added to a 100 mL beaker and stirred until the pyridoxal phosphate was completely dissolved to obtain a homogeneous solution. 50 mL of the solution was transferred from the beaker to a 200 mL reaction flask, and 1 g of the compound and 0.1 g of aminotransferase containing mutant 231 were added to the solution. The mixture was vortexed and incubated at 47 °C for 20 h. After the reaction, the solution was removed, and 10 mL of water was added. Extraction was performed with 200 mL of methyl tert-butyl ether (pH adjusted to 13 with potassium hydroxide). After extraction, the organic phase was removed, and anhydrous sodium sulfate was added to remove water. The filtrate was then filtered and distilled under reduced pressure at -0.09 MPa (gauge pressure). The product was purified by column chromatography, yielding 0.749 g of the product (98.3%). NMR analysis showed that the purity of the chiral lactam was greater than 98%.

[0348] Example 8

[0349] In sodium borate buffer, aminotransferase catalyzes the synthesis of lactam from compound 1.

[0350]

[0351] Add 0.06 g of pyridoxal phosphate and 90 mL of 0.05 mol / L sodium borate buffer (pH 10.5) containing isopropylamine to a 100 mL beaker, and stir until the pyridoxal phosphate is completely dissolved to obtain a homogeneous solution. Take 28 mL of the solution from the beaker and add it to a 100 mL reaction flask, then add 1 g of the compound and 0.05 g of aminotransferase containing mutant 440 to the solution. Vortex to mix the resulting solution and react at 47 °C for 20 h. After the reaction was completed, the reaction solution was removed, and 10 mL of water was added. Extraction was then performed using 200 mL of methyl tert-butyl ether (pH adjusted to 13 with potassium hydroxide). Following extraction, the organic phase was removed, and anhydrous sodium sulfate was added to remove water. The filtrate was then filtered and distilled under reduced pressure at -0.09 MPa (gauge pressure). Purification was then performed by column chromatography, yielding 0.621 g of the product, with a yield of 99.1%. NMR analysis showed that the purity of the chiral lactam was greater than 98%.

[0352] Example 9

[0353] In sodium borate buffer, aminotransferase catalyzes the synthesis of lactam from compound 5.

[0354]

[0355] 0.06 g of pyridoxal phosphate and 90 mL of 0.05 mol / L sodium borate buffer (pH 9.0) containing isopropylamine were added to a 100 mL beaker and stirred until the pyridoxal phosphate was completely dissolved to obtain a homogeneous solution. 50 mL of the solution was transferred from the beaker to a 200 mL reaction flask, and 1 g of the compound and 0.1 g of aminotransferase containing mutant 73 were added to the solution. The mixture was vortexed and incubated at 47 °C for 20 h. After the reaction, the solution was removed, and 10 mL of water was added. Extraction was performed with 200 mL of methyl tert-butyl ether (pH adjusted to 13 with potassium hydroxide). After extraction, the organic phase was removed, and anhydrous sodium sulfate was added to remove water. The filtrate was then filtered and distilled under reduced pressure at -0.09 MPa (gauge pressure). The product was purified by column chromatography, yielding 0.720 g of the product (94.5%). NMR analysis showed that the purity of the chiral lactam was greater than 98%.

[0356] Example 10

[0357] In sodium borate buffer, aminotransferase catalyzes the synthesis of lactam from compound 3.

[0358]

[0359] 0.06 g of pyridoxal phosphate and 90 mL of 0.05 mol / L sodium borate buffer (pH 10.5) containing isopropylamine were added to a 100 mL beaker. The mixture was stirred until all pyridoxal phosphate was dissolved to obtain a homogeneous solution. 28 mL of the solution was transferred from the beaker to a 100 mL reaction flask, and 1 g of the compound and 0.1 g of aminotransferase containing mutant 344 were added. The solution was vortexed and mixed, and then reacted at 37 °C for 20 h. After the reaction, the reaction solution was removed, and 10 mL of water was added. The mixture was then extracted with 200 mL of methyl tert-butyl ether (pH adjusted to 13 with potassium hydroxide). After extraction, the organic phase was removed, and anhydrous sodium sulfate was added to remove water. The filtrate was then filtered and distilled under reduced pressure at -0.09 MPa (gauge pressure). The product was purified by column chromatography, yielding 0.65 g of the product (98.9%). NMR analysis showed that the purity of the chiral lactam was greater than 98%.

[0360] Example 11

[0361] In sodium borate buffer, aminotransferase catalyzes the synthesis of lactam from compound 2.

[0362]

[0363] 0.06 g of pyridoxal phosphate and 90 mL of 0.05 mol / L sodium borate buffer (pH 10) containing isopropylamine were added to a 100 mL beaker. The mixture was stirred until pyridoxal phosphate was completely dissolved to obtain a homogeneous solution. 28 mL of the solution was transferred from the beaker to a 100 mL reaction flask, and 1 g of the compound and 0.05 g of aminotransferase containing mutant 280 were added. The mixture was vortexed to mix the solution and then reacted at 47 °C for 20 h. After the reaction, the reaction solution was removed, and 10 mL of water was added. The mixture was then extracted with 200 mL of methyl tert-butyl ether (pH adjusted to 13 with potassium hydroxide). After extraction, the organic phase was removed, and anhydrous sodium sulfate was added to remove water. The filtrate was then filtered and distilled under reduced pressure at -0.09 MPa (gauge pressure). The product was purified by column chromatography, yielding 0.645 g of the product, with a yield of 98.1%. NMR analysis showed that the purity of the chiral lactam was greater than 98%.

[0364] Example 12

[0365] In sodium borate buffer, aminotransferase catalyzes the synthesis of lactam from compound 6.

[0366]

[0367] 0.06 g of pyridoxal phosphate and 90 mL of 0.05 mol / L sodium borate buffer (pH 10) containing isopropylamine were added to a 100 mL beaker. The mixture was stirred until pyridoxal phosphate was completely dissolved to obtain a homogeneous solution. 50 mL of the solution was transferred from the beaker to a 200 mL reaction flask, and 1 g of the compound and 0.1 g of aminotransferase containing mutant 368 were added. The mixture was vortexed to mix the solution and then reacted at 47 °C for 20 h. After the reaction, the reaction solution was removed, and 10 mL of water was added. The mixture was then extracted with 200 mL of methyl tert-butyl ether (pH adjusted to 13 with potassium hydroxide). After extraction, the organic phase was removed, and anhydrous sodium sulfate was added to remove water. The filtrate was then filtered and distilled under reduced pressure at -0.09 MPa (gauge pressure). The product was purified by column chromatography, yielding 0.572 g of the product (99.3%). NMR analysis showed that the purity of the chiral lactam was greater than 98%.

[0368] Example 13

[0369] In sodium borate buffer, aminotransferase catalyzes the synthesis of lactam from compound 7.

[0370]

[0371] 0.06 g of pyridoxal phosphate and 90 mL of 0.05 mol / L sodium borate buffer (pH 10.5) containing isopropylamine were added to a 100 mL beaker. The mixture was stirred until pyridoxal phosphate was completely dissolved to obtain a homogeneous solution. 28 mL of the solution was transferred from the beaker to a 100 mL reaction flask, and 1 g of the compound and 0.1 g of aminotransferase containing mutant 398 were added. The solution was vortexed and mixed, and then reacted at 55 °C for 20 h. After the reaction, the reaction solution was removed, and 10 mL of water was added. The mixture was then extracted with 200 mL of methyl tert-butyl ether (pH adjusted to 13 with potassium hydroxide). After extraction, the organic phase was removed, and anhydrous sodium sulfate was added to remove water. The filtrate was then filtered and distilled under reduced pressure at -0.09 MPa (gauge pressure). The product was purified by column chromatography, yielding 0.743 g of the product (92.4% yield). NMR analysis showed that the purity of the chiral lactam was greater than 98%.

[0372] Example 14

[0373] In sodium borate buffer, aminotransferase catalyzes the synthesis of lactam from compound 7.

[0374]

[0375] Add 0.06 g of pyridoxal phosphate and 90 mL of 0.05 mol / L sodium borate buffer (pH 10.5) containing isopropylamine to a 100 mL beaker, and stir until the pyridoxal phosphate is completely dissolved to obtain a homogeneous solution. Transfer 50 mL of the solution from the beaker to a 200 mL reaction flask, and add 1 g of the compound and 0.05 g of aminotransferase containing mutant 423 to the solution. Vortex to mix the resulting solution and react at 47 °C for 20 h. After the reaction was completed, the reaction solution was removed, and 10 mL of water was added. Extraction was then performed using 200 mL of methyl tert-butyl ether (pH adjusted to 13 with potassium hydroxide). Following extraction, the organic phase was removed, and anhydrous sodium sulfate was added to remove water. The filtrate was then filtered and distilled under reduced pressure at -0.09 MPa (gauge pressure). Purification was then performed by column chromatography, yielding 0.797 g of the product, with a yield of 99.1%. NMR analysis showed that the purity of the chiral lactam was greater than 98%.

[0376] Example 15

[0377] In potassium phosphate buffer, aminotransferase catalyzes the synthesis of lactam from compound 1.

[0378]

[0379] 0.06 g of pyridoxal phosphate and 90 mL of 0.05 mol / L sodium borate buffer (pH 10.5) containing isopropylamine were added to a 100 mL beaker and stirred until the pyridoxal phosphate was completely dissolved to obtain a homogeneous solution. 28 mL of the solution was transferred from the beaker to a 100 mL reaction flask, and 1 g of the compound and 0.1 g of aminotransferase with mutant 256 were added to the solution. The mixture was vortexed and incubated at 47 °C for 20 h. After the reaction, the solution was removed, 10 mL of water was added, and the mixture was extracted with 200 mL of methyl tert-butyl ether (pH adjusted to 13 with potassium hydroxide). After extraction, the organic phase was removed, and anhydrous sodium sulfate was added to remove water. The filtrate was then filtered and distilled under reduced pressure at -0.09 MPa (gauge pressure). The filtrate was purified by column chromatography to obtain 0.622 g of the product, with a yield of 99.4%. NMR analysis showed that the purity of the chiral lactam was greater than 98%.

[0380] Example 16

[0381] In potassium phosphate buffer, aminotransferase catalyzes the synthesis of lactam from compound 2.

[0382]

[0383] 0.06 g of pyridoxal phosphate and 90 mL of 0.05 mol / L sodium borate buffer (pH 10.5) containing isopropylamine were added to a 100 mL beaker and stirred until the pyridoxal phosphate was completely dissolved to obtain a homogeneous solution. 28 mL of the solution was transferred from the beaker to a 100 mL reaction flask, and 1 g of the compound and 0.1 g of aminotransferase with mutant 49 were added to the solution. The mixture was vortexed and incubated at 37 °C for 20 h. After the reaction, the solution was removed, 10 mL of water was added, and the mixture was extracted with 200 mL of methyl tert-butyl ether (pH adjusted to 13 with potassium hydroxide). After extraction, the organic phase was removed, and anhydrous sodium sulfate was added to remove water. The filtrate was then filtered and distilled under reduced pressure at -0.09 MPa (gauge pressure). The filtrate was purified by column chromatography to obtain 0.643 g of the product, with a yield of 97.9%. NMR analysis showed that the purity of the chiral lactam was greater than 98%.

[0384] Example 17

[0385] In potassium phosphate buffer, aminotransferase catalyzes the synthesis of lactam from compound 3.

[0386]

[0387] 0.06 g of pyridoxal phosphate and 90 mL of 0.05 mol / L sodium borate buffer (pH 9.5) containing isopropylamine were added to a 100 mL beaker and stirred until the pyridoxal phosphate was completely dissolved to obtain a homogeneous solution. 50 mL of the solution was transferred from the beaker to a 200 mL reaction flask, and 1 g of the compound and 0.1 g of aminotransferase with mutant 356 were added to the solution. The mixture was vortexed and incubated at 47 °C for 20 h. After the reaction, the solution was removed, 10 mL of water was added, and the mixture was extracted with 200 mL of methyl tert-butyl ether (pH adjusted to 13 with potassium hydroxide). After extraction, the organic phase was removed, and anhydrous sodium sulfate was added to remove water. The filtrate was then filtered and distilled under reduced pressure at -0.09 MPa (gauge pressure). The product was purified by column chromatography, yielding 0.646 g of the product (98.3%). NMR analysis showed that the purity of the chiral lactam was greater than 98%.

[0388] Example 18

[0389] In Tris solution, aminotransferase catalyzes the synthesis of lactam from compound 2.

[0390]

[0391] 0.06 g of pyridoxal phosphate and 90 mL of 0.05 mol / L Tris buffer (pH 10.5) were added to a 100 mL beaker and stirred until the pyridoxal phosphate was completely dissolved to obtain a homogeneous solution. 50 mL of the solution was transferred from the beaker to a 200 mL reaction flask, and 1 g of the compound and 0.1 g of aminotransferase with mutant 418 were added to the solution. The mixture was vortexed and incubated at 27 °C for 20 h. After the reaction, the solution was removed, 10 mL of water was added, and the mixture was extracted with 200 mL of methyl tert-butyl ether (pH adjusted to 13 with potassium hydroxide). After extraction, the organic phase was removed, and anhydrous sodium sulfate was added to remove water. The filtrate was then filtered and distilled under reduced pressure at -0.09 MPa (gauge pressure). The product was purified by column chromatography, yielding 0.614 g of the product (93.4% yield). NMR analysis showed that the purity of the chiral lactam was greater than 98%.

[0392] Example 19

[0393] In Tris solution, aminotransferase catalyzes the synthesis of lactam from compound 4.

[0394]

[0395] 0.06 g of pyridoxal phosphate and 90 mL of 0.05 mol / L Tris buffer (pH 10.5) were added to a 100 mL beaker and stirred until the pyridoxal phosphate was completely dissolved to obtain a homogeneous solution. 50 mL of the solution was transferred from the beaker to a 200 mL reaction flask, and 1 g of the compound and 0.1 g of aminotransferase with mutant 18 were added to the solution. The mixture was vortexed and incubated at 47 °C for 20 h. After the reaction, the solution was removed, 10 mL of water was added, and the mixture was extracted with 200 mL of methyl tert-butyl ether (pH adjusted to 13 with potassium hydroxide). After extraction, the organic phase was removed, and anhydrous sodium sulfate was added to remove water. The filtrate was then filtered and distilled under reduced pressure at -0.09 MPa (gauge pressure). The product was purified by column chromatography, yielding 0.643 g of the product (97.9%). NMR analysis showed that the purity of the chiral lactam was greater than 98%.

[0396] Example 20

[0397] In TEOA buffer, aminotransferase catalyzes the synthesis of lactam from compound 7.

[0398]

[0399] 0.06 g of pyridoxal phosphate and 90 mL of 0.05 mol / L TEOA buffer (pH 10.5) were added to a 100 mL beaker and stirred until the pyridoxal phosphate was completely dissolved to obtain a homogeneous solution. 28 mL of the solution was transferred from the beaker to a 100 mL reaction flask, and 1 g of the compound and 0.1 g of aminotransferase with mutant 406 were added to the solution. The mixture was vortexed and incubated at 47 °C for 20 h. After the reaction, the solution was removed, 10 mL of water was added, and the mixture was extracted with 200 mL of methyl tert-butyl ether (pH adjusted to 13 with potassium hydroxide). After extraction, the organic phase was removed, and anhydrous sodium sulfate was added to remove water. The filtrate was then filtered and distilled under reduced pressure at -0.09 MPa (gauge pressure). The product was purified by column chromatography, yielding 0.790 g of the product (98.3%). NMR analysis showed that the purity of the chiral lactam was greater than 98%.

[0400] Example 21

[0401] In TEOA solution, aminotransferase catalyzes the synthesis of lactam from compound 8.

[0402]

[0403] 0.06 g of pyridoxal phosphate and 90 mL of 0.05 mol / L TEOA sodium buffer (pH 11) were added to a 100 mL beaker and stirred until the pyridoxal phosphate was completely dissolved to obtain a homogeneous solution. 28 mL of the solution was transferred from the beaker to a 100 mL reaction flask, and 1 g of the compound and 0.1 g of aminotransferase with mutant 441 were added to the solution. The mixture was vortexed and incubated at 47 °C for 20 h. After the reaction, the solution was removed, 10 mL of water was added, and the mixture was extracted with 200 mL of methyl tert-butyl ether (pH adjusted to 13 with potassium hydroxide). After extraction, the organic phase was removed, and anhydrous sodium sulfate was added to remove water. The filtrate was then filtered and distilled under reduced pressure at -0.09 MPa (gauge pressure). The product was purified by column chromatography, yielding 0.800 g of the product (94.1%). NMR analysis showed that the purity of the chiral lactam was greater than 98%.

[0404] Example 22

[0405] In TEOA solution, aminotransferase catalyzes the synthesis of lactam from compound 6.

[0406]

[0407] 0.06 g of pyridoxal phosphate and 90 mL of 0.05 mol / L TEOA buffer (pH 10) were added to a 100 mL beaker and stirred until the pyridoxal phosphate was completely dissolved to obtain a homogeneous solution. 50 mL of the solution was transferred from the beaker to a 200 mL reaction flask, and 1 g of the compound and 0.1 g of aminotransferase with mutant 245 were added to the solution. The mixture was vortexed and incubated at 47 °C for 20 h. After the reaction, the solution was removed, 10 mL of water was added, and the mixture was extracted with 200 mL of methyl tert-butyl ether (pH adjusted to 13 with potassium hydroxide). After extraction, the organic phase was removed, and anhydrous sodium sulfate was added to remove water. The filtrate was then filtered and distilled under reduced pressure at -0.09 MPa (gauge pressure). The product was purified by column chromatography, yielding 0.568 g of the product (98.7%). NMR analysis showed that the purity of the chiral lactam was greater than 98%.

[0408] Example 23

[0409] The specific mutation sites and types are shown in Table 1. The conditions are the same as in Example 8, except that the mutant of the catalytic enzyme used in the reaction is replaced with the mutant in Table 1. The reaction results are shown in Table 2.

[0410] Table 2

[0411]

[0412]

[0413]

[0414]

[0415]

[0416]

[0417]

[0418]

[0419]

[0420]

[0421]

[0422]

[0423]

[0424]

[0425]

[0426]

[0427] The reaction conditions and effects of the examples are shown in Tables 3-1 and 3-2 below:

[0428] Table 3-1

[0429]

[0430]

[0431] Table 3-2

[0432]

[0433] The sequence information used in this invention is as follows:

[0434] The amino acid sequence of the wild-type aminotransferase is: MSLTVQKINWEQVKEWDRKYLMRTFSTQNEYQPVPIESTEGDYLIMPDGTRLLDFFNQLYCVNLGQKNQKVNAAIKEALDRYGFVWDTYATDYKAKAAKIIIEDILGDEDWPGKVRFVSTGSEAVETALNIARLYTNRPLVVTREHDYHGWTGGAATVTRLRSYRSGLVGENSESFSAQIPGSSYNSAVLMAPSPNMFQDSDGNLLKDENGELLSVKYTRRMIENYGPEQV AAVITEVSQGAGSAMPPYEYIPQIRKMTKELGVLWINDEVLTGFGRTGKWFGYQHYGVQPDIITMGKGLSSSSLPAGAVLVSKEIAAFMDKHRWESVSTYAGHPVAMAAVCANLEVMMEENFV EQAKDSGEYIRSKLELLQEKHKSIGNFDGYGLLWIVDIVNAKTKTPYVKLDRNFTHGMNPNQIPTQIIMKKALEKGVLIGGVMPNTMRIGASLNVSRGDIDKAMDALDYALDYLESGEWQ(SEQ ID NO:1)

[0435]

[0436] The above embodiments include preferred embodiments of the present invention, but the embodiments of the present invention are not limited to the above embodiments. Any changes, modifications, substitutions, combinations, or simplifications made without departing from the spirit and principle of the present invention shall be considered equivalent substitutions and shall be included within the protection scope of the present invention.

Claims

1. An aminotransferase mutant, characterized in that, Compared to the amino acid sequence shown in SEQ ID NO:1, it contains differences in one or more of the following amino acid residues: position 7, position 8, position 9, position 11, position 12, position 15, position 16, position 18, position 19, position 20, position 21, position 22, position 24, position 25, position 28, position 29, position 32, position 34, position 37, position 40, position 48, position 51, position 56, position 58, position 59, position 60, position 61, position 67, position 69, position 70, position 73, position 76, position 77, position 79, position 80, position 81, position 84, position 86, position 87, position 88, position 89, position 90, position 96. 99th, 103rd, 104th, 108th, 110th, 114th, 119th, 120th, 121st, 122nd, 123rd, 124th, 125th, 134th, 135th, 137th, 148th, 149th, 150th, 151st, 152nd 155th, 156th, 158th, 159th, 161st, 162nd, 163rd, 164th, 165th, 168th, 170th, 171st, 172nd, 174th, 175th, 176th, 177th, 178th, 179th, 181st, 1st 82nd, 185th, 187th, 191st, 197th, 198th, 199th, 200th, 201st, 202nd, 204th, 205th, 207th, 209th, 210th, 217th, 218th, 221st, 224th, 225th, 226th 229th, 237th, 240th, 241st, 242nd, 243rd, 244th, 245th, 249th, 250th, 257th, 260th, 261st, 269th, 271st, 272nd, 273rd, 280th, 286th, 288th, 290th 295th, 297th, 307th, 314th, 315th, 321st, 322nd, 323rd, 324th, 325th, 326th, 327th, 328th, 329th, 330th, 331st, 332nd, 337th, 346th, 349th, 3rd 50th, 351st, 352nd, 355th, 356th, 358th, 359th, 362nd, 365th, 369th, 370th, 372nd, 373rd, 374th, 376th, 380th, 387th, 396th, 397th, 398th, 399thPositions 400, 401, 403, 404, 408, 415, 416, 420, 423, 424, 427, 428, 429, 436, 437, 442, 448, 451, 456, 459, 463, 466, 467, 469, 472, and 474; and the aminotransferase mutant exhibits activity superior to that of the aminotransferase with the amino acid sequence shown in SEQ ID NO:

1.

2. The aminotransferase mutant as described in claim 1, characterized in that, Compared with the amino acid sequence shown in SEQ ID NO:1, it contains one or more differences selected from the following amino acid residues: (1) the 7th position is: A, C, D, F, G, H, I, L, M, N, P, Q, S, T, V, W or Y; (2) the 8th position is: C, D, E, F, H, K, L, M, N, Q, R, T, V, W or Y; (3) the 9th position is: A, C, D, E, G, H, I, K, L, R, S, T, V, W or Y; (4) the 11th position is: A, C, D, F, G, H, I, K, L, M, N, P, Q, R, S, V or Y; (5) the 12th position is: A, C, D, E, F, G, H, I, L, M, P, S, T, V, W or Y; (6) The 15th digit is: C, D, F, H, I, K, L, M, N, P, Q, R, S, W or Y; (7) The 16th digit is: A, E, F, G, I, K, L, M, N, Q, R or S; (8) The 18th digit is: C, D, E, G, I, M, N, P, Q, S, T or V; (9) The 19th digit is: A, C, D, E, F, G, H, M, N, P, S, T, W or Y; (10) The 20th digit is: A, C, D, G, I, K, L, M, N, P, Q, R, S, T or V; (11) The 21st digit is: A, C, D, F, G, H, I, K, M, P, Q, R, T, V, W or Y; (12) The 22nd digit is: A, C, D (13) The 24th bit is: A, C, D, E, F, G, H, K, L, N, Q, R, S, V, W or Y; (14) The 25th bit is: A, C, D, E, F, G, H, K, P, Q, R, S, T, V or Y; (15) The 28th bit is: C, D, E, F, H, I, K, L, M, P, R, S, T or Y; (16) The 29th bit is: C, D, F, L, P, Q, R, S or Y; (17) The 32nd bit is: A, C, D, E, F, G, H, K, L, M, P, R, S, T, V, W or Y; (18) The 34th bit is: C, D, E, F, G, H (19) The 37th bit is: F, G, H, I, K, L, P, Q, R, S, V or W; (20) The 40th bit is: A, C, G, H, I, K, L, N, P, R, S, T, W or Y; (21) The 48th bit is: C, D, E, F, G, M, N, Q, R, S, T, V, W or Y; (22) The 51st bit is: C, D, G, H, I, K, L, M, Q, R, T, V, W or Y; (23) The 56th bit is: A, C, D, F, G, I, L, N, P, R, S, V or Y; (24) The 58th bit is: D, E, F, G, H, I, K, N, R, S, T, V, W or Y;(25) The 59th digit is: C, D, E, F, G, I, K, N, P, Q, R, S, T, V, W or Y; (26) The 60th digit is: A, C, D, G, H, I, K, F, M, N, P, Q, R, S, T or W; (27) The 61st digit is: A, D, E, F, G, H, I, K, L, M, P, Q, R, V or W (28) The 67th digit is: A, C, D, E, G, H, I, L, M, N, P, Q, R, S, T, W or Y; (29) The 69th digit is: A, D, F, H, I, K, M, N, P, R, S, T, W or Y; (30) The 70th digit is: A, C, D, E, F, H, I, L, M, P, Q, R, S, T or W; (31) The 73rd digit is: C, E, F, G, H, I, K, L, M, N, Q, R, S, V, W or Y; (32) The 76th digit is: A, C, D, E, F, G, H, I, L, N, P, Q, R, S, T, V or W; (33) The 77th digit is: A, C, D, F, G, H, K, L, M, N, P, Q, R, S Or Y; (34) The 79th bit is: C, D, E, G, I, K, N, P, Q, R, S, T, V, W or Y; (35) The 80th bit is: A, C, E, F, G, H, I, K, L, M, N, P, R, T, V, W or Y; (36) The 81st bit is: C, E, I, K, L, N, P, Q, S, T, V, W or Y; 37) The 84th digit is: A, C, D, E, H, I, K, L, N, P, R, T, V, W, or Y; (38) The 86th digit is: A, C, D, E, F, G, I, K, L, M, N, P, Q, R, S, T, V, or Y; (39) The 87th digit is: A, C, E, F, G, H, K, L, M, N, P, Q, R, S, T, V, W or Y; (40) The 88th bit is: A, C, D, E, F, G, H, I, K, L, M, N, P, Q, R, V, W or Y; (41) The 89th bit is: A, D, E, F, G, H, I, L, P, S, T, V or W; (42) The 90th bit is: C, E, F, G, H, K, L, P, Q, S, T, V (43) The 96th bit is: A, C, D, E, G, H, I, L, N, P, Q, R, S, T, V, W or Y; (44) The 99th bit is: A, D, F, G, H, I, N, P, Q, R, S, T, V, W or Y; (45) The 103rd bit is: A, D, F, G, H, I, K, L, P, Q, R, S (46) The 104th bit is: A, C, E, F, H, I, L, M, N, Q, R, V or W; (47) The 108th bit is: A, C, E, G, H, I, K, M, P, R, S, T, V or W; (48) The 110th bit is: A, C, F, G, I, K, L, M, P, Q, R, T, V, W or Y;(49) The 114th digit is: A, C, D, E, F, G, I, L, M, N, P, Q, R, S, V, W, or Y; (50) The 119th digit is: A, C, D, E, F, G, H, L, M, N, Q, R, V, W, or Y; (51) The 120th digit is: A, C, D, E, F, G, H, I, K, L, M, N, Q, R, S, W, or Y; (52) The 121st digit is: D, E, F, H, I, M, N, P, R, S, V, or Y; (53) The 122nd digit is: A, D, F, G, H, I, K, L, M, P, T, V, W, or Y; (54) The 123rd digit is: A, D, F, G, H, I, K, L, M, P, Q, R, S, V, or W; (55) The 124th bit is: C, D, E, G, I, K, L, N, P, Q, S, W, or Y; (56) The 125th bit is: A, C, D, F, G, H, I, L, M, N, Q, S, T, W, or Y; (57) The 134th bit is: A, D, E, F, H, I, K, M, P, Q, S, T, or W; (58) The 135th bit is: A, C, D, E, F, H, I, L, N, P, Q, R, S, T, V, or W; (59) The 137th bit is: A, D, F, G, H, I, K, L, M, P, R, T, or W; (60) The 148th bit is: C, D, E, F, G, H, I, K, L, M, N, P, S, T, V, W or Y; (61) The 149th bit is: A, C, D, E, F, G, I, K, L, M, P, R, S, T, V or W; (62) The 150th bit is: A, C, D, F, H, I, K, L, M, N, Q, R, S, T, W or Y; (63) The 151st bit is: A, C, D, G, H, K, L, M, N, Q, R, S, T, V or Y; (64) The 152nd bit is: A, C, D, E, F, G, H, K, L, M, N, P, Q, R, S, V, W or Y; (65) The 155th bit is: C, E, F, I, L, M, N, Q, R, V or Y; (66) The 156th bit is: C, D, E, F, G, H, I, K, L, M, N, P, Q, R, S, T, V or W; (67) the 158th bit is: A, C, D, E, F, G, H, I, K, L, M, N, P, R, S, T, W or Y; (68) the 159th bit is: C, D, F, G, H, K, L, N, Q, R, S, V, W or Y; (69) the 161st bit is: A, D, E, G, I, K, M, P, Q, R, S, T, V, W or Y; (70) the 162nd bit is: A, C, D, E, F, G, H, I, K, L, N, P, Q, S, T, V, W or Y; (71) the 163rd bit is: A, C, D, G, H, I, L, M, N, P, R, T, W or Y;(72) The 164th digit is: A, C, D, E, F, G, H, I, K, L, M, N, P, Q, R, S, T, V, or W; (73) The 165th digit is: A, C, F, G, H, I, K, L, M, P, Q, R, V, W, or Y; (74) The 168th digit is: A, C, D, E, F, G, H, I, K, N, P, Q (75) The 170th bit is: A, C, D, E, I, K, L, M, N, P, Q, R, S, V, W, or Y; (76) The 171st bit is: A, D, E, F, H, I, K, L, M, N, P, R, S, T, V, or Y; (77) The 172nd bit is: A, C, D, E, F, H, I, K, M, P, Q, R, S, T, V, W or Y; (78) The 174th bit is: A, D, F, G, H, I, L, M, N, P, Q, R, S, V, W or Y; (79) The 175th bit is: A, C, D, E, F, G, H, I, K, L, M, N, P, Q, R, T, V, W or Y; (80) The 176th bit is: A, C, D (81) The 177th bit is: A, C, D, E, F, G, H, I, K, L, M, N, P, Q, T, V, W or Y; (82) The 178th bit is: C, D, F, G, H, I, K, L, N, P, Q, R, T, V, W or Y; (83) The 179th digit is: C, D, F, G, H, I, L, N, Q, R, S, T, V, W, or Y; (84) The 181st digit is: D, E, F, G, H, I, K, M, Q, S, or V; (85) The 182nd digit is: A, C, E, F, H, I, L, P, Q, S, T, V, or Y; (86) The 185th digit is: A, C, D, E, F, H, K, L, M, N, P, Q, R, S, T, V or W; (87) The 187th bit is: A, C, E, F, G, H, L, M, N, P, Q, R, T or Y; (88) The 191st bit is: A, C, D, E, F, G, H, I, K, N, P, Q, S, T, V or W; (89) The 197th bit is: A, D, E, F, G, H, I, K, L, N, P, Q, R, S, T, V, W or Y; (90) the 198th position is: A, C, E, G, H, I, K, M, P, Q, T, V, W or Y; (91) the 199th position is: A, C, D, E, F, G, H, I, K, L, M, N, P, R, S, T, W or Y; (92) the 2nd position is: A, C, D, E, F, G, H, I, K, L, M, N, P, R, S, T, W or Y; The 00th digit is: A, C, E, F, H, K, L, M, N, Q, R, T, V, W or Y; (93) The 201st digit is: A, C, D, E, F, G, H, I, K, L, M, N, P, Q, R, T, V, W or Y; (94) The 202nd digit is: A, C, E, F, G, H, K, L, M, P, R, S, T, W or Y;(95) The 204th digit is: A, C, D, F, G, H, I, K, L, Q, R, T, W, or Y; (96) The 205th digit is: A, C, D, E, F, G, H, I, K, M, N, P, Q, R, S, T, W, or Y; (97) The 207th digit is: A, E, G, I, M, N, P, Q, R, S, V, W, or Y; 98) The 209th digit is: A, D, F, G, I, K, N, Q, R, T, V, W or Y; (99) The 210th digit is: A, D, E, F, G, H, I, K, M, P, R, T or V; (100) The 217th digit is: A, C, D, E, F, G, H, I, M, N, P, Q, R, S, T, V, W or Y; (10) 1) The 218th bit is: A, D, E, F, G, H, K, L, M, N, P, S or T; (102) The 221st bit is: A, C, D, E, F, H, I, L, M, N, P, Q, S, T, V, W or Y; (103) The 224th bit is: A, C, D, F, G, H, K, L, P, Q, R, S, T or V; (10 4) The 225th bit is: A, C, D, E, F, G, H, K, L, M, P, Q, S, T, V, W or Y; (105) The 226th bit is: A, C, D, F, H, I, K, L, M, P, Q, R, S, T or V; (106) The 229th bit is: A, C, D, F, G, H, I, K, N, P, Q, R, S, T (107) The 237th bit is: A, C, D, F, G, H, I, K, L, M, N, P, S, T, V or W; (108) The 240th bit is: A, E, F, G, H, K, L, M, P, S, V, W or Y; (109) The 241st bit is: A, D, H, I, K, P, R, S, T, V or Y; 110) The 242nd bit is: C, D, F, G, H, I, K, L, N, S, T, V, W or Y; (111) The 243rd bit is: A, C, D, F, H, I, K, N, P, Q, S, T, V or W; (112) The 244th bit is: A, C, E, F, G, H, I, K, L, N, Q, V, W or Y; (113) The 245th digit is: C, D, E, F, G, H, I, K, L, M, N, P, R, S, T, V, W, or Y; (114) The 249th digit is: A, C, D, E, G, H, I, K, L, M, N, P, Q, R, S, T, V, or W; (115) The 250th digit is: A, C, D, G, H, I, M, Q, R, S, or T (116) The 257th bit is: A, D, E, F, H, L, M, N, P, R, S, T, V, W or Y; (117) The 260th bit is: C, D, E, F, G, H, L, N, P, Q, R, S, T or Y; (118) The 261st bit is: A, C, D, F, G, H, K, N, P, Q, R, S, T, W or Y;(119) The 269th bit is: A, C, E, F, I, K, L, M, N, P, Q, S, T, V, W, or Y; (120) The 271st bit is: C, D, E, F, G, H, K, L, M, S, T, or Y; (121) The 272nd bit is: A, C, E, G, P, Q, R, T, W, or Y; (122) The 273rd bit is: D, E, F, G, H, I, K, L, M, N, P, Q, R, S, or Y; (123) The 280th bit is: C, F, G, H, I, L, M, N, P, Q, S, T, V, or W; (124) The 286th bit is: A, C, E, F, G, I, K, N, Q, R, S, T, V, W, or Y; (125) The 288th bit is: A, C, D, E, I, K, M, N, Q, R, S, V, W or Y; (126) The 290th bit is: A, C, D, E, F, G, H, I, K, L, M, P, R, S, T, V, W or Y; (127) The 295th bit is: A, C, D, E, F, G, H, I, K, L, M, N, P, Q, S, V or Y; (128) The 297th bit is: A, C, D, E, K, L, M, N, Q, R, S, V or W; (129) The 307th bit is: C, D, E, F, G, H, I, K, L, M, N, P, Q, S, T, V or W; (130) The 314th bit is: A, D, E, F, G, H, I, L, M (131) The 315th bit is: C, D, F, G, H, I, K, M, N, Q, R, S, T, V, W or Y; (132) The 321st bit is: A, C, E, F, G, H, I, K, L, M, N, P, Q, R, S, T, V or Y; (133) The 322nd bit is: A, C, E, F, G, H, I, M, N, P, R, S, T, V or W; (134) The 323rd bit is: A, C, D, E, F, G, K, L, N, P, Q, R, V, W or Y; (135) The 324th bit is: C, F, G, I, K, L, M, N, P, Q, S or Y; (136) The 325th bit is: C, E, F, H, I, K, L, M, N, P, Q, S, T, V or Y; (137) the 326th bit is: C, F, G, H, K, L, M, P, R, S, T or V; (138) the 327th bit is: C, D, E, F, G, H, K, L, M, N, P, Q, R, T, V, W or Y; (139) the 328th bit is: A, C, D, E, F, G, H, I, K, L, M, P, Q, R, S, T, W or Y; (140) the 329th bit is: A, C, E, G, I, L, N, P, Q, R, T, V, W or Y; (141) the 330th bit is: A, C, D, E, F, G, H, I, L, M, P, Q, S, V or W;(142) The 331st bit is: A, C, D, E, F, G, H, I, K, L, M, N, P, Q, R, S, T, V, or W; (143) The 332nd bit is: D, E, F, H, I, K, N, P, Q, R, S, T, W, or Y; (144) The 337th bit is: C, D, E, F, G, H, I, K, L, M, N, P, Q, R, S, T, V, W, or Y; (145) The 346th bit is: A, C, D, F, G, H, I, K, N, P, Q, T, V, or Y; (146) The 349th bit is: D, E, F, H, I, L, N, P, Q, T, V, W, or Y; (147) The 350th bit is: C, D, F, G, I, K, L, M, P, Q, R, S, T, V, W or Y; (148) the 351st bit is: A, C, F, H, I, K, N, P, Q, R, S, T or W; (149) the 352nd bit is: C, D, E, F, H, I, K, L, P, Q, R, S, T, V or W; (150) the 355th bit is: A, C, H, I, L, N, P, Q, R, T, W or Y; (151) the 356th bit is: A, D, E, G, H, I, N, Q, S, T, V, W or Y; (152) the 358th bit is: C, E, F, H, I, M, N, P, R, S, T, V or W; (153) the 359th bit is: A, E, F, G, H, I, K, N, P, Q (154) The 362nd bit is: A, C, F, G, H, I, K, M, N, P, R, S, T, or Y; (155) The 365th bit is: A, C, D, E, F, G, H, I, L, N, P, Q, S, T, V, W, or Y; (156) The 369th bit is: D, F, K, L, M, N, Q, R, S, T, V, W, or Y; (157) The 370th bit is: C, D, E, F, G, I, K, M, N, P, Q, R, S, T, V, W, or Y; (158) The 372nd bit is: A, C, D, E, F, G, I, K, L, M, N, P, R, S, T, V, W, or Y; (159) The 373rd bit is: (160) The 374th bit is: A, C, D, F, G, H, I, L, M, N, P, Q, V, or Y; (161) The 376th bit is: D, E, G, I, M, N, Q, R, T, W, or Y; (162) The 380th bit is: A, C, D, E, F, G, H, I, K, L, M, P, Q, S, T, V, or Y; (163) The 387th bit is: A, C, D, E, F, G, H, I, K, M, N, P, Q, R, S, T, V, W, or Y; (164) The 396th bit is: A, C, E, F, G, H, I, L, M, N, P, Q, S, T, W, or Y;(165) The 397th digit is: A, C, D, E, F, G, H, I, K, M, N, R, S, V, or W; (166) The 398th digit is: A, C, D, E, F, G, H, I, L, M, N, P, R, T, V, W, or Y; (167) The 399th digit is: A, C, E, F, H, L, M, N, P, Q, R, S (168) The 400th bit is: A, C, D, E, F, G, H, I, L, M, N, R, S, W or Y; (169) The 401st bit is: A, D, E, F, H, I, K, L, N, P, S, T or W; (170) The 403rd bit is: A, C, D, E, F, G, H, I, M, N, P, Q, R, S, T, V, W or Y; (171) the 404th bit is: A, C, D, G, K, M, N, R, T, V, W or Y; (172) the 408th bit is: A, C, D, E, G, H, I, K, L, M, N, P, Q, R, S, T or V; (173) the 415th bit is: A, C, D, E, F, G, H, I, K (174) The 416th bit is: A, C, D, F, G, H, I, K, L, M, N, P, R, S, T, or W; (175) The 420th bit is: A, D, E, G, H, K, L, P, R, S, W, or Y; (176) The 423rd bit is: A, C, F, G, I, K, L, M, N (177) The 424th bit is: D, E, F, G, L, M, N, P, Q, R, S, V, W, or Y; (178) The 427th bit is: A, C, D, E, F, G, H, I, K, M, N, P, R, S, T, V, W, or Y; (179) The 428th bit is: C, D, F, G, K, L, M, N, P, R, or T; (180) the 429th bit is: A, C, D, F, G, H, I, L, M, N, P, Q, R, S, T, V, W, or Y; (181) the 436th bit is: A, C, D, E, F, G, H, I, K, L, N, P, Q, R, S, T, W, or Y; (182) the 437th bit is: A C, D, E, F, G, H, I, L, N, P, Q, R, S, T or Y; (183) The 442nd bit is: A, C, E, F, G, H, I, L, M, N, Q, S, T, V, W or Y; (184) The 448th bit is: A, C, D, E, F, G, H, I, K, L, M, P, Q, R, S, T or Y; (1 85) The 451st bit is: A, C, D, E, H, I, K, M, N, P, Q, S, T, V, W or Y; (186) The 456th bit is: D, E, F, G, H, I, L, N, P, Q, R, S, T, V or Y; (187) The 459th bit is: A, C, E, F, G, L, M, N, P, R, S, V, W or Y;(188) The 463rd bit is: A, C, D, E, F, G, H, I, L, M, N, P, Q, R, S, T, V, or W; (189) The 466th bit is: A, C, E, F, G, H, I, K, L, M, N, P, Q, R, T, V, or Y; (190) The 467th bit is: A, C, E, F, G, H, I, K, L, M, N, Q, R, S, T or V; (191) The 469th bit is: A, C, D, F, H, I, K, L, M, N, P, S, T, V, W or Y; (192) The 472nd bit is: A, C, D, G, H, K, L, M, N, P, S, T, W or Y; (193) The 474th bit is: A, C, D, E, F, G, H, I, K, N, P, R, S, T, V, W or Y.

3. The aminotransferase mutant as described in claim 1, characterized in that, It satisfies any of the following conditions: (1) The aminotransferase mutant, compared with the amino acid sequence shown in SEQ ID NO:1, contains amino acid residue differences at the following sites: 007D and 073N, and position 060 is F or M; preferably, the aminotransferase mutant also contains differences at one or more of the following sites: 076R, 088I, 096R, 156R, 158T, 162K, 164M and 170A; the aminotransferase mutant may also contain differences at one or more of the following sites: 175E, 178Q, 197R, 199R, 201G, 205Y, 221D, 245T, 249R, 290H, 295I and 327T; (2) The aminotransferase mutant, compared with the amino acid sequence shown in SEQ ID NO:1, contains amino acid residue differences at the following sites: 007D and 073N, and position 060 is F or M; Compared with the amino acid sequence shown in SEQ ID NO:1, the aminotransferase mutant contains amino acid residue differences at one or more of the following sites: 086W, 175E, 199T, 201G, 221D, 249R, 328I, 370D, 380S, 387M, 436T, 448N, I or R at position 088, M or L at position 164, and W or F at position 331; (3) Compared with the amino acid sequence shown in SEQ ID NO:1, the aminotransferase mutant contains amino acid residue differences at the following sites: 007D, 021I, 073N, 076R, 096R, 156R, 158T, 162K, 170A, 178Q, and 197R; preferably, the aminotransferase mutant also contains amino acid residue differences at the following sites: 205Y, 245T, 290H, 295I, 327T, and 337C; more preferably, the aminotransferase mutant also contains The aminotransferase mutant contains the following differences in amino acid residues: 372S, 403P, 408A, and 442I; the aminotransferase mutant may also contain one or more differences in amino acid residues at the following sites: 060 is S, M, or F; 088 is V, I, H, N, or R; 199 is R or T; 211 is A or D; 249 is K or R; 328 is M or I; 380 is G or S; 436 is L or T; (4) the aminotransferase mutant is similar to that in SEQ Compared to the amino acid sequence shown in ID NO:1, the amino acid residues at the following sites differ: 007D, 021I, 060 is S or F or M, 073N, 076R, 088 is V or I or N or H or R, 096R, 156R, 158T, 162K, 170A, 178Q, 197R, 199 is R or T, 205Y, 221 is A or D, 245T, 249 is K or R, 290H, 295I, 327T, 328 is M or I, 337C, 372S, 380 is G or S, 403P, 408A, 436 is L or T and 442I;Preferably, it also includes amino acid differences at one or more of the following sites: 086T, M or L at position 164, 175E, 176W, 177A, 201G, F or W at position 331, D or E at position 370, 387M, and 448A.

4. The aminotransferase mutant as described in claim 1, characterized in that, The aminotransferase mutant, compared with the amino acid sequence shown in SEQ ID NO:1, has any of the following combinations of amino acid residue differences:

5. An enzyme preparation comprising the aminotransferase mutant according to any one of claims 1-4; preferably, the enzyme preparation is a buffer solution containing the aminotransferase mutant; the buffer solution is, for example, sodium borate buffer, phosphate buffer, trimethylolpropionate buffer or triethanolamine buffer; more preferably, the enzyme preparation is a powder, such as a lyophilized powder.

6. An isolated polynucleotide, said polynucleotide encoding an aminotransferase mutant as described in any one of claims 1-4.

7. A nucleic acid construct comprising the polynucleotide as described in claim 6; preferably, the nucleic acid construct is an expression cassette or vector, such as a cloning vector or expression vector.

8. A transformant comprising the polynucleotide of claim 6 or the nucleic acid construct of claim 7; preferably, the host cell used in constructing the transformant is Escherichia coli.

9. A method for preparing an aminotransferase mutant as described in any one of claims 1-4, characterized in that, The method includes: culturing the transformant as described in claim 8 to obtain the aminotransferase mutant.

10. A method for preparing a lactam compound, characterized in that, It includes the following steps: In the presence of the aminotransferase mutant as described in any one of claims 1-4, the compound shown in Formula I and the amino donor are subjected to an amination reaction to obtain a lactam compound. R3 is an oxo group, -CHO, or... Y1 and Y2 are independently H or -SO2X, where X is H or an alkali metal; R1 and R2 are independently C. 1-6 Alkyl, C 6-14 aryl or C substituted with one or more R4 6-14 Aryl; R4 independently is C 1-6 Alkyl, C 1-6 alkoxy, halogen, or hydroxyl; when R3 is an oxo group or When the lactam compound is When R3 is -CHO, the lactam compound is And * indicates that C is in the S configuration; preferably, the preparation method of the lactam compound satisfies one or more of the following conditions: (1) R3 is an oxo group or -CHO; (2) R2 is C 1-6 Alkyl; (3) R3 is C 1-6 Alkyl or aryl; (4) Y1 is H, Y2 is -SO2X, X is H or an alkali metal, such as sodium; more preferably, the compound shown in Formula I is or, for or, for 11. The method for preparing a lactam compound according to claim 10, characterized in that, It satisfies one or more of the following conditions: (1) the amino donor is an amine compound or its salt, such as isopropylamine, L-alanine, DL-alanine, L-lysine, DL-lysine, (S)-1-phenylethylamine, (R)-1-phenylethylamine, (RS)-1-phenylethylamine, pentanediamine, o-phenylenediamine, o-phenylenediamine hydrochloride, 2-(4-nitrobenzene)ethylamine, and 2-(4-nitrobenzene)ethylamine hydrochloride; preferably, the amino donor is isopropylamine; (2) the molar ratio of the amino donor to the compound of Formula I is (1-6):1, for example 4:1; (3) the reaction solvent for the amination reaction is a buffer solution or a mixture of a buffer solution and an organic solvent, wherein the organic solvent Examples of suitable solvents include methyl tert-butyl ether, isopropyl ether, tetrahydrofuran, 2-methyltetrahydrofuran, 1,4-dioxane, ethyl acetate, dimethyl sulfoxide, N,N-dimethylformamide, methanol, ethanol, isopropanol, or n-butanol. When the reaction solvent for the amination reaction is a mixture of a buffer solution and an organic solvent, the organic solvent is preferably present in a volume ratio of 0 to 250 mL / L. More preferably, the reaction solvent for the amination reaction is a buffer solution, such as one or more of sodium borate buffer, phosphate buffer, trimethylolpropane buffer, and triethanolamine buffer. Even more preferably, the volume-to-mass ratio of the buffer solution to the compound of Formula I is 5 to 150 mL / g, for example, 20 mL / g. -110 mL / g, preferably 28 mL / g, 50 mL / g or 100 mL / g; (4) the pH value of the amination reaction is 9.0 to 11; preferably 9.0 to 10.5; for example 9.0, 9.5, 10.0, 10.5 or 11.0; (5) the reaction temperature of the amination reaction is 25℃ to 50℃; preferably 47℃, 37℃, 55℃ or 27℃; (6) the mass ratio of the aminotransferase mutant to the compound shown in Formula I is (0.01-1):1, for example 0.05:1 or 0.1:1; (7) the amination reaction includes the following steps: adding the compound shown in Formula I and the aminotransferase mutant to a mixture containing the buffer solution and the amino donor, After mixing evenly, the reaction is carried out at 25℃~50℃; (8) After the amination reaction is completed, the following post-treatment steps are also included: after the reaction is completed, the pH is adjusted, water and ether solvent are added for extraction and separation, the organic phase is dried and the ether solvent is removed; preferably, the post-treatment steps satisfy one or more of the following conditions a, b, c and d: condition a: the ether solvent is tert-butyl methyl ether; condition b: the drying is drying with a desiccant; the desiccant is, for example, anhydrous sodium sulfate; condition c: the pH is adjusted by using an alkali, the alkali is preferably sodium hydroxide or potassium hydroxide, for example, potassium hydroxide; condition d: the ether solvent is removed by vacuum distillation, for example, vacuum distillation at a pressure of -0.09Mpa.

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