Transaminase mutant and application thereof

A transaminase and mutant technology, applied in the field of transaminase mutants and their applications, can solve the problem that transaminase is difficult to catalyze the synthesis of large-sterically hindered chiral amine compounds

Inactive Publication Date: 2021-03-16
ASYMCHEM LAB TIANJIN
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] The main purpose of the present invention is to provide a transaminase mutant and its application to solve the problem

Method used

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  • Transaminase mutant and application thereof
  • Transaminase mutant and application thereof
  • Transaminase mutant and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0076] Directed mutations were carried out on the basis of the mother parent. The specific mutation sites are shown in the table below, and the catalytic activity of the mutants was tested according to the following reaction conditions:

[0077] 1 mL system includes 1 mg substrate 1, 1 mg PLP, 1 mg isopropylamine hydrochloride, 50 mg enzyme powder, pH8.0 100mM phosphate buffer, react for 40 h.

[0078] The test results are shown in the table below:

[0079] Table 1:

[0080]

[0081] .

[0082] Note: In the above table, 0 means the conversion rate is <1%, + means the conversion rate is greater than or equal to 1% and less than 5%, and ++ means the conversion rate is greater than or equal to 5% and less than or equal to 10%.

Embodiment 2

[0084] On the basis of Example 1, the combined mutation was carried out, and the activity of the combined mutation was screened according to the same reaction conditions as in Example 1. The results are shown in the table below.

[0085] Table 2:

[0086]

[0087] .

[0088] Note: In the above table, 0 means the conversion rate is <1%, + means the conversion rate is greater than or equal to 1% and less than 5%, ++ means the conversion rate is greater than or equal to 5% and less than 10%, +++ means the conversion rate is greater than or equal to 10 % and less than 15%, ++++ means the conversion rate is greater than or equal to 15% and less than 20%.

Embodiment 3

[0090] On the basis of Example 2, combined mutations were carried out, and the activity of combined mutations was screened according to the same reaction conditions as in Example 1. The results are shown in the table below.

[0091] table 3:

[0092]

[0093] .

[0094] Note: 0 means the conversion rate is <1%, + means the conversion rate is greater than or equal to 1% and less than 5%, ++ means the conversion rate is greater than or equal to 5% and less than 10%, +++ means the conversion rate is greater than or equal to 10% and less than 15% %, ++++ means the conversion rate is greater than or equal to 15% and less than 20%, and +++++ means the conversion rate is greater than or equal to 20% and less than or equal to 40%.

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Abstract

The invention provides a transaminase mutant and an application thereof. The transaminase mutant is obtained by mutation of one or more amino acids in SEQ ID NO: 2 or is a mutant which is obtained bymutation of conservative amino acids at the 54th-63rd site and the 84th-96th site by taking the sequence SEQ ID NO: 1 of wild type CvTA transaminase as a reference. The catalytic activity of the mutants is improved to different extents compared with that of wild transaminase, so that mutants are used for synthesizing chiral amine compounds, especially synthesizing chiral amine with large steric hindrance, the production efficiency is improved, and the industrial production cost is reduced. The biological enzyme is adopted for green chemical synthesis, thereby being beneficial to reducing industrial three wastes.

Description

technical field [0001] The invention relates to the application field of biological enzymes, in particular to a transaminase mutant and its application. Background technique [0002] Large hindered chiral amine compounds (in this application, refers to a class of compounds where the group next to the potential chiral carbonyl group is larger than the methyl group) are a class of optically active substances and functional molecules, and are important compounds widely used in the synthesis of drugs and ligands. chiral intermediates. However, there are few reports on the synthesis of such chiral compounds, and satisfactory results cannot be obtained. For example, the heavily hindered phenyl tert-butyl amino ester on the carbonyl side has a selectivity of 76% (Angew. Chem. Int. Ed. 2007, 46, 4367). At present, some metal catalysts, such as ruthenium, rhodium, palladium, etc., have been reported for the asymmetric hydrogenation reaction of small steric hindrance (in this applic...

Claims

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Application Information

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IPC IPC(8): C12N9/10C12N15/54C12P13/00
CPCC12N9/1096C12Y206/01C12P13/001C12P17/12C12P17/14C12N15/52
Inventor 洪浩詹姆斯·盖吉肖毅张娜焦学成马玉磊牟慧艳王祖建孙凯华李响赵桐曹珊
Owner ASYMCHEM LAB TIANJIN
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