Glucosyltransferase amino acid motifs for enzymatic production of linear poly alpha-1,3-glucan
Reactions comprising water, sucrose, and one or more glucosyltransferase enzymes are disclosed herein. Glucosyltransferase enzymes used in these reactions comprise certain motifs allowing production of insoluble poly alpha-1,3-glucan having at least 95% alpha-1,3 glycosidic linkages.
1. A method for identifying a glucosyltransferase enzyme, wherein said identifying consists of:
detecting the presence of at least one motif in a glucosyltransferase catalytic domain, wherein said at least one motif is selected from the group consisting of:
a motif comprising an amino acid sequence that is at least 90% identical to SEQ ID NO:78,
(ii) a motif comprising an amino acid sequence that is at least 90% identical to SEQ ID NO:79, and
(iii) a motif comprising an amino acid sequence that is at least 90% identical to SEQ ID NO:80;
thereby identifying a glucosyltransferase enzyme that produces insoluble poly alpha-1,3-glucan having at least 95% alpha-1,3 glycosidic linkages and a weight average degree of polymerization (DP w ) of at least 100.
2. The method of claim 1 , wherein the detecting step is performed:
(a) in silico,
(b) with a method comprising a nucleic acid hybridization step,
(c) with a method comprising a protein sequencing step, and/or
(d) with a method comprising a protein binding step.
3. The method of claim 1 , wherein said identifying consists of detecting the presence of each of motifs (i), (ii) and (iii) in said catalytic domain.
4. The method of claim 1 , further comprising, after said identifying, producing the glucosyltransferase enzyme.
5. The method of claim 4 , wherein the glucosyltransferase enzyme is produced by a method comprising recombinant expression of the glucosyltransferase enzyme.
6. The method of claim 5 , wherein said recombinant expression comprises microbial expression of the glucosyltransferase enzyme.
7. The method of claim 4 , further comprising:
contacting the glucosyltransferase enzyme with at least water and sucrose, thereby producing said insoluble poly alpha-1,3-glucan.
8. The method of claim 1 , wherein the catalytic domain comprises an amino acid sequence that is at least 90% identical to amino acid positions 54-957 of SEQ ID NO:65.
9. The method of claim 8 , wherein the catalytic domain comprises an amino acid sequence that is at least 95% identical to amino acid positions 54-957 of SEQ ID NO:65.
10. The method of claim 8 , wherein
(A) the position of the amino acid sequence that is at least 90% identical to SEQ ID NO:78 aligns with amino acid positions 231-243 of SEQ ID NO:65;
(B) the position of the amino acid sequence that is at least 90% identical to SEQ ID NO:79 aligns with amino acid positions 396-425 of SEQ ID NO:65; and/or
(C) the position of the amino acid sequence that is at least 90% identical to SEQ ID NO:80 aligns with amino acid positions 549-567 of SEQ ID NO:65.
11. The method of claim 1 , wherein the glucosyltransferase enzyme produces poly alpha-1,3-glucan having at least 99% alpha-1,3 glycosidic linkages.
12. The method of claim 1 , wherein the glucosyltransferase enzyme produces poly alpha-1,3-glucan having a DP w of at least 400.
13. The method of claim 1 , wherein the glucosyltransferase enzyme produces poly alpha-1,3-glucan having 100% alpha-1,3 glycosidic linkages.
14. The method of claim 7 , wherein said insoluble poly alpha-1,3-glucan has at least 99% alpha-1,3 glycosidic linkages.
15. The method of claim 7 , wherein said insoluble poly alpha-1,3-glucan has 100% alpha-1,3 glycosidic linkages.