Modified alpha-glucosidase and applications of same

Patent metadata

Field	Value
Publication number	US-9650619-B2
Application number	US-201615278556-A
Country	US
Kind code	B2
Filing date	Sep 28, 2016
Priority date	Mar 16, 2011
Publication date	May 16, 2017
Grant date	May 16, 2017

Abstract

The object is to provide an α-glucosidase in which a transglycosylation activity predominates, and use thereof, and the like. A modified α-glucosidase consisting of an amino acid sequence in which one or two or more of the amino acid(s) is selected from a group of specific amino acids.

First claim

The invention claimed is:

1. A modified α-glucosidase which (a) consists of an amino acid sequence which is identical to SEQ ID NO: 2 except for a substitution at a position corresponding to position 495 of SEQ ID NO: 2, or (b) comprises an amino acid sequence which has 95% or more identity with SEQ ID NO: 2 and comprises a substitution at a position corresponding to position 495 of SEQ ID NO: 2,
wherein the amino acid at position 495 is substituted by a alanine, glycine, proline, threonine or valine, and
wherein the modified α-glucosidase has increased transglycosylation activity to form α-1,6 glucosidic linkages compared to the α-glucosidase of SEQ ID NO: 2.

2. A method for producing an oligosaccharide, comprising reacting the modified α-glucosidase of claim 1 with an oligosaccharide or polysaccharide having an α-1,4 glucosidic linkage.

3. A method for producing an oligosaccharide, comprising reacting the modified α-glucosidase of claim 1 with an oligosaccharide or polysaccharide having α-1,6 glucosidic linkage.

4. The modified α-glucosidase of claim 1 , wherein said modified α-glucosidase comprises an amino acid sequence which has 95% or more identity with SEQ ID NO: 2 and comprises a substitution at a position corresponding to position 495 of SEQ ID NO: 2, wherein the amino acid at position 495 is substituted by a alanine, glycine, proline, threonine or valine, wherein the modified α-glucosidase has increased transglycosylation activity to form α-1,6 glucosidic linkages compared to the α-glucosidase of SEQ ID NO: 2, and wherein said modified α-glucosidase further comprises one or more amino acid substitutions at an amino acid selected from the group consisting of:
(i) an amino acid corresponding to the amino acid at position 343 of SEQ ID NO: 2;
(ii) an amino acid corresponding to the amino acid at position 452 of SEQ ID NO: 2;
(iii) an amino acid corresponding to the amino acid at position 496 of SEQ ID NO: 2;
(iv) an amino acid corresponding to the amino acid at position 410 of SEQ ID NO: 2;
(v) an amino acid corresponding to the amino acid at position 498 of SEQ ID NO: 2;
(vi) an amino acid corresponding to the amino acid at position 499 of SEQ ID NO: 2;
(vii) an amino acid corresponding to the amino acid at position 531 of SEQ ID NO: 2;
(viii) an amino acid corresponding to the amino acid at position 533 of SEQ ID NO: 2;
(ix) an amino acid corresponding to the amino acid at position 579 of SEQ ID NO: 2;
(x) an amino acid corresponding to the amino acid at position 585 of SEQ ID NO: 2;
(xi) an amino acid corresponding to the amino acid at position 662 of SEQ ID NO: 2;
(xii) an amino acid corresponding to the amino acid at position 715 of SEQ ID NO: 2; and
(xiii) an amino acid corresponding to the amino acid at position 721 of SEQ ID NO: 2.

Assignees

• Amano Enzyme Inc

Inventors

• Ishihara Satoru

Classifications

• C12P19/18

  produced by the action of a glycosyl transferase, e.g. alpha-, beta- or gamma-cyclodextrins · CPC title

• C12N9/2408 Primary

  acting on alpha -1,4-glucosidic bonds · CPC title

• Y02P20/52

  using catalysts, e.g. selective catalysts · CPC title

• C12N9/2428

  Glucan 1,4-alpha-glucosidase (3.2.1.3), i.e. glucoamylase · CPC title

• C12N9/1051

  Hexosyltransferases (2.4.1) · CPC title