A group of cyclodextrin glucosyltransferase, and coding gene and application thereof

A cyclodextrin and gene technology, applied in the direction of transferase, application, genetic engineering, etc., can solve the problem of low specificity of CGTases, and achieve the effect of high specificity and high production efficiency

Active Publication Date: 2012-12-19
INST OF MICROBIOLOGY - CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The specificity of CGTases is low, and in most cases the thre

Method used

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  • A group of cyclodextrin glucosyltransferase, and coding gene and application thereof
  • A group of cyclodextrin glucosyltransferase, and coding gene and application thereof
  • A group of cyclodextrin glucosyltransferase, and coding gene and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0067] Embodiment 1, the discovery of mutated protein and its coding gene

[0068] A strain of Bacillus macerans screened from nature can produce cyclodextrin glycosyltransferase, and the cyclodextrin glycosyltransferase gene in the strain is shown in sequence 2 of the sequence listing, encoding sequence 1 of the sequence listing proteins indicated.

[0069] The protein shown in Sequence 1 is named α-CGTase-1 protein, and the gene encoding it is named α-CGTase-1 gene (as shown in Sequence 2). The protein shown in Sequence 3 was named α-CGTase-WT protein (sequence published in NCBI), and the gene encoding it was named α-CGTase-WT gene (shown in Sequence 4).

[0070] The difference between Sequence 1 and Sequence 3 lies only in the 590th amino acid residue (M / V) and the 677th amino acid residue (S / G). The difference between sequence 2 and sequence 4 lies in the 1768th nucleotide (A / G) and the 2029th nucleotide (A / G).

[0071] Using the double-stranded DNA shown in Sequence 1 ...

Embodiment 2

[0072] Embodiment 2, enzyme activity identification

[0073] 1. Artificially synthesize the following double-stranded DNA molecules:

[0074] (1) Double-stranded DNA molecule 1: The difference from the DNA molecule shown in Sequence 2 of the Sequence Listing is that nucleotides 499-501 from the 5' end are mutated from TAC to TGG, and nucleotides 1606-501 from the 5' end The 1608th nucleotide is mutated from GCG to GTG; correspondingly, the 167th amino acid residue in sequence 1 is mutated from tyrosine to tryptophan, and the 536th amino acid residue is mutated from alanine to valine acid. The protein encoded by double-stranded DNA molecule 1 is named protein 1.

[0075] (2) Double-stranded DNA molecule 2: The difference from the DNA molecule shown in Sequence 2 of the Sequence Listing is that nucleotides 499-501 from the 5' end are mutated from TAC to TGC, and nucleotides 1606-501 from the 5' end The 1608th nucleotide is mutated from GCG to GTG; correspondingly, the 167th a...

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Abstract

The invention discloses a group of cyclodextrin glucosyltransferase, and a coding gene and an application thereof. The cyclodextrin glucosyltransferase provided by the invention comes from Bacillus macerans, and is a protein obtained by mutating a protein as shown in a sequence 1 of a sequence table through the following two mutations: (a) mutating an amino acid residue at 536th site of N-end of the sequence 1 in the sequence table from alanine to valine, and (b) mutating an amino acid residue at 167th site of N-end of the sequence 1 in the sequence table. The protein provided by the invention is relatively high in specificity of producing alpha-CD, relatively high in production efficiency, and has great value to production.

Description

technical field [0001] The invention relates to a group of cyclodextrin glucosyltransferases and their coding genes and applications. Background technique [0002] The English name of cyclodextrin is cyclodextrin, referred to as CD. Cyclodextrin is the product of cyclodextrin glucosyltransferase (CGTase) acting on starch. It is a cyclic oligosaccharide with more than six glucose linked by α-1,4-glycosidic bonds. The most common and most studied is α-cyclodextrin (α-CD), β-cyclodextrin (β-CD), γ-cyclodextrin (γ-CD), respectively composed of six, seven and eight glucose molecules, are relatively large and relatively flexible molecules. [0003] α-cyclodextrin is a kind of high-value product of starch deep processing, which is a cyclic oligosaccharide composed of six glucose groups linked by α-1,4 glucosidic bonds. In terms of structure, the three-dimensional structure of α-CD is cylindrical (one end is large and the other end is small). It has the particularity of "hydrophi...

Claims

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Application Information

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IPC IPC(8): C12N9/10C12N15/54C12N15/63C12N5/10C12N1/21C12P19/18C12R1/19
Inventor 钞亚鹏钱世钧宋炳红
Owner INST OF MICROBIOLOGY - CHINESE ACAD OF SCI
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