Method for detecting D-2-hydroxyglutarate by using FAD dependent D-2-hydroxyglutarate dehydrogenase and resazurin
A technology of hydroxyglutarate dehydrogenase and hydroxyglutarate, which is applied in botany equipment and methods, biochemical equipment and methods, enzymes, etc., can solve the problem of poor substrate specificity, high detection cost, and sensitivity low cost, low cost, high sensitivity and low cost
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[0071] 1. Preparation of AX-F-D2HGDH protein
[0072] 1.1 Acquisition of gene sequence
[0073] Achromobacter xylosoxidansATCC27061 is a common model strain of Achromobacter, and the A-F-D2HGDH derived from this Achromobacter is named AX-F-D2HGDH.
[0074] Enter Gene ID: 29509945 into the NCBI database to obtain the gene sequence (SEQ ID NO: 1, 1413bp) and amino acid sequence (SEQ ID NO: 2, 470aa) of AX-F-D2HGDH in Achromobacter xylosoxidansATCC27061.
[0075] Using sequence alignment tools well-known to researchers in the field (such as Nucleotide Blast in NCBI, etc.), the gene sequence of AX-F-D2HGDH was compared to obtain a sequence with an identity greater than 70%, and the encoded protein had D-2- Hydroxyglutarate dehydrogenase activity. Using sequence alignment tools well known to researchers in the field (such as Protein Blast in NCBI, etc.), compare the amino acid sequence of AX-F-D2HGDH to obtain a sequence with more than 50% identity, and the protein has D-2-hydrox...
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[0128] 1. Preparation of AX-F-D2HGDH protein
[0129] 1.1 Acquisition of gene sequence
[0130] Achromobacter xylosoxidansATCC27061 is a common model strain of Achromobacter, and the A-F-D2HGDH derived from this Achromobacter is named AX-F-D2HGDH.
[0131]Enter Gene ID: 29509945 into the NCBI database to obtain the gene sequence (SEQ ID NO: 1, 1413bp) and amino acid sequence (SEQ ID NO: 2, 470aa) of AX-F-D2HGDH in Achromobacter xylosoxidansATCC27061.
[0132] Using sequence alignment tools well-known to researchers in the field (such as Nucleotide Blast in NCBI, etc.), the gene sequence of AX-F-D2HGDH was compared to obtain a sequence with an identity greater than 70%, and the encoded protein had D-2- Hydroxyglutarate dehydrogenase activity, and has high substrate specificity, only active on D-2-HG. Utilize the sequence alignment tools well-known to researchers in the field (such as ProteinBlast in NCBI, etc.), compare the amino acid sequence of AX-F-D2HGDH, and obtain a seq...
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