METHOD FOR PRODUCING ALPHA-1, 6-BRANCHED ALPHA-1, 4-GLUCANS FROM SUCROSE

• US 20080020432A1
• Filed: 07/20/2007
• Published: 01/24/2008
• Est. Priority Date: 10/09/1998
• Status: Active Grant

• Current Assignee: Bayer Cropscience AG (Bayer AG), Max-Planck-Gesellschaft Zur Foerderung Der Wissenschaften e.V. (Max-Planck-Gesellschaft zur Frderung der Wissenschaften e)
• Original Assignee: Bayer Bioscience GmbH, Max-Planck-Gesellschaft Zur Foerderung Der Wissenschaften e.V. (Max-Planck-Gesellschaft zur Frderung der Wissenschaften e)
• Inventors: Quanz, Martin, Buttcher, Volker
• Granted Patent: US 7,638,311 B2
• Time in Patent Office: Days
• US Class Current: 435/101
• CPC Class Codes: C07K 14/22 from Neisseriaceae (F)
• CPC Class Codes: C12N 9/1051 Hexosyltransferases (2.4.1)
• CPC Class Codes: C12N 9/107 1,4-Alpha-glucan branching enzyme (2.4.1.18)
• CPC Class Codes: C12P 19/04 Polysaccharides, i.e. compounds containing more than five saccharide radicals attached to each other by glycosidic bonds
• CPC Class Codes: C12P 19/16 produced by the action of an alpha-1, 6-glucosidase, e.g. amylose, debranched amylopectin non-biological hydrolysis of starch C08B30/00
• CPC Class Codes: C12P 19/20 produced by the action of an exo-1,4 alpha-glucosidase, e.g. dextrose

Abstract

Nucleic acid molecules which encode a branching enzyme from a bacterium of the genus Neisseria, vectors, host cell, plant cells and plants containing said nucleic acid molecules as well as starch obtainable from the plants described are described. Furthermore, an in-vitro method for producing α-1,6-branched α-1,4-glucans on the basis of sucrose and a combination of enzymes of an amylosucrase and a branching enzyme as well as the α-1,6-branched α-1,4-glucans obtainable by said method are described.

Claims

1. An α

-1,6-branched α

-1,4-glucan, wherein said glucan has a branching degree that is higher than glycogen.

2. An α

-1,6-branched α

-1,4-glucan according to claim 1, wherein said glucan has a molecular weight in the range of 1000-3000×

10⁶ kd.

3. An α

-1,6-branched α

-1,4-glucan according to claim 1, wherein said glucan has a molecular weight in the range of 100,000-1500×

10⁶ kd.

4. An α

-1,6-branched α

-1,4-glucan according to claim 1, wherein said glucan has a molecular weight in the range of 100,000-1000×

10⁶ kd.

5. An α

-1,6-branched α

-1,4-glucan according to claim 1, wherein said glucan has a molecular weight in the range of 262,000-1000×

10⁶ kd.

6. An α

-1,6-branched α

-1,4-glucan according to claim 1, wherein said glucan has a molecular weight in the range of 262,000-499×

10⁶ kd.

13. A method for producing α

- 1,6-branched α

-1,4-glucans in in-vitro systems comprising

(1) expressing in an in-vitro system a protein encoded by a nucleic acid molecule encoding a branching enzyme from a bacterium of the genus Neisseria selected from the group consisting of

(a) nucleic acid molecules encoding a protein which comprises the amino acid sequence depicted in SEQ ID NO. 2;

(b) nucleic acid molecules comprising the coding region depicted in SEQ ID NO. 1;

(c) nucleic acid molecules encoding a protein which comprises the amino acid sequence encoded by the insert in plasmid DSM 12425;

(d) nucleic acid molecules comprising the coding region for a branching enzyme, which is contained in the insert of the plasmid DSM 12425;

(e) nucleic acid molecules encoding a protein the sequence of which has, in the first 100 amino acids, a homology of at least 65% to the amino acid sequence depicted in SEQ ID NO. 2;

(f) nucleic acid molecules the complementary strand of which hybridizes to a nucleic acid molecule of (a), (b), (c), (d) and/or (e) and which encode a branching enzyme from a bacterium of the genus Neisseria; and

(g) nucleic acid molecules the sequence of which deviates from the sequence of a nucleic acid molecule of (f) due to the degeneracy of the genetic code;

or

obtainable by a method according to claim 1;

(2) contacting said protein with sucrose and amylosucrase; and

(3) isolating said α

-1,6-branched α

-1,4-glucans.

7. An α

-1,6-branched α

-1,4-glucan, wherein said glucan has a branching degree that is higher than glycogen, said glucan produced by a method comprising

(a) identifying the activity of an amylosucrase and a branching enzyme;

(b) mixing said amylosucrase and said branching enzyme in a ratio of protein activity of 1;

4000-2000;

1;

(c) reacting said mixture of amylosucrase and branching enzyme with a sucrose solution; and

(d) isolating an α

-1,6-branched α

-1,4-glucan with a branching degree that is higher than glycogen.

8. An α

-1,6-branched α

-1,4-glucan according to claim 7, wherein said glucan has a molecular weight in the range of 1000-3000×

10⁶ kd.

9. An α

-1,6-branched α

-1,4-glucan according to claim 7, wherein said glucan has a molecular weight in the range of 100,000-1500×

10⁶ kd.

10. An α

-1,6-branched α

-1,4-glucan according to claim 7, wherein said glucan has a molecular weight in the range of 100,000-1000×

10⁶ kd.

11. An α

-1,6-branched α

-1,4-glucan according to claim 7, wherein said glucan has a molecular weight in the range of 262,000-1000×

10⁶ kd.

12. An α

-1,6-branched α

-1,4-glucan according to claim 7, wherein said glucan has a molecular weight in the range of 262,000-499×

10⁶ kd.