Glucosyltransferase enzymes for production of glucan polymers
View Patent ↗Reaction solutions are disclosed herein comprising water, sucrose and a glucosyltransferase enzyme that synthesizes poly alpha-1,3-glucan. The glucosyltransferase enzyme can synthesize insoluble glucan polymer having at least 50% alpha-1,3 glycosidic linkages and a number average degree of polymerization of at least 100. Further disclosed are methods of using such glucosyltransferase enzymes to produce insoluble poly alpha-1,3-glucan.
1. A method for producing insoluble poly alpha-1,3-glucan comprising:
a) contacting at least water, sucrose, and an isolated glucosyltransferase enzyme comprising an amino acid sequence that is at least 90% identical to SEQ ID NO:4, whereby the glucose transferase enzyme uses sucrose as a substrate to produce said insoluble poly alpha-1,3-glucan having at least 90% alpha-1,3 glycosidic linkages; and
b) isolating the insoluble poly alpha-1,3-glucan produced in step (a).
2. The method of claim 1 , wherein said insoluble poly alpha-1,3-glucan has a number average degree of polymerization of at least 100.
3. The method of claim 1 , wherein said insoluble poly alpha-1,3-glucan has at least 95% alpha-1,3 glycosidic linkages.
4. The method of claim 3 , wherein said insoluble poly alpha-1,3-glucan has at least 97% alpha-1,3 glycosidic linkages.
5. The method of claim 4 , wherein said insoluble poly alpha-1,3-glucan has at least 99% alpha-1,3 glycosidic linkages.
6. The method of claim 5 , wherein said insoluble poly alpha-1,3-glucan has about 100% alpha-1,3 glycosidic linkages.
7. The method of claim 1 , wherein step (a) further comprises contacting a primer with the water, sucrose, and glucosyltransferase enzyme.
8. The method of claim 7 , wherein the primer comprises dextran.
9. The method of claim 1 , wherein said glucosyltransferase enzyme comprises an amino acid sequence that is at least 93% identical to SEQ ID NO:4.
10. The method of claim 1 , wherein said glucosyltransferase enzyme comprises an amino acid sequence that is at least 95% identical to SEQ ID NO:4.
11. The method of claim 1 , wherein said glucosyltransferase enzyme comprises an amino acid sequence that is at least 97% identical to SEQ ID NO:4.
12. The method of claim 1 , wherein said glucosyltransferase enzyme comprises an amino acid sequence that is at least 98% identical to SEQ ID NO:4.
13. The method of claim 1 , wherein said glucosyltransferase enzyme comprises an amino acid sequence that is at least 99% identical to SEQ ID NO:4.
14. The method of claim 1 , wherein said glucosyltransferase enzyme does not comprise amino acid residues 2-1477 or 138-1477 of SEQ ID NO:8.
15. The method of claim 1 , wherein a heterologous amino acid sequence of 1-300 residues is at the N-terminus and/or C-terminus of said glucosyltransferase enzyme.
16. The method of claim 1 , wherein step (a) comprises only one of said glucosyltransferase enzyme.
17. The method of claim 1 , wherein the water, sucrose, and glucosyltransferase enzyme are contacted in cell-free conditions.
18. The method of claim 1 , wherein step (b) of isolating the poly alpha-1,3-glucan comprises filtration.
19. The method of claim 1 , wherein step (b) of isolating the poly alpha-1,3-glucan comprises centrifugation.
20. The method of claim 1 , wherein the poly alpha-1,3-glucan isolated in step (b) is washed with water.