High titer production of poly (α 1,3 glucan)
View Patent ↗A process for enzymatic preparation of poly (α1, 3 glucan) from sucrose is disclosed. The glucosyltransferase enzyme (gtfJ) from Streptococcus salivarius is used to convert sucrose to fructose and poly (α1, 3 glucan). Application of semi-permeable membranes to continuously remove fructose, a by-product of the gtf enzyme, thus increasing the poly (α1, 3 glucan) liter, is disclosed.
1. A reaction system for producing poly (alpha 1,3 glucan), wherein said reaction system comprises:
a) a first chamber that comprises an enzyme reaction solution comprising:
i) sucrose, and
ii) at least one glucosyltransferase enzyme that is a Streptococcus salivarius GtfJ enzyme; and
b) a second chamber, separated from the first chamber by a semi-permeable membrane that is in contact with the enzyme reaction solution, wherein the semi-permeable membrane is permeable to fructose and sucrose, but impermeable to poly (alpha 1,3 glucan) and the glucosyltransferase enzyme, and wherein the semi-permeable membrane facilitates continuous removal of fructose and other low molecular weight moieties from the first chamber while retaining poly (alpha 1,3 glucan) and the at least one glucosyltransferase enzyme inside the first chamber, wherein the semi-permeable membrane has a molecular weight cut-off from 12,000 to 100,000 Daltons.
2. The reaction system of claim 1 , wherein said enzyme reaction solution further comprises at least one primer.
3. The reaction system of claim 1 , wherein the glucosyltransferase enzyme is a primer-independent enzyme.
4. The reaction system of claim 1 , wherein the semi-permeable membrane facilitates accumulation of poly (alpha 1,3 glucan) in the first chamber to a concentration ranging from 30 grams per liter to 200 grams per liter.
5. The reaction system of claim 4 , wherein the semi-permeable membrane is a dialysis tubing.
6. The reaction system of claim 2 , wherein the at least one primer is dextran.
7. The reaction system of claim 1 , wherein more than one glucosyltransferase enzyme is present in the enzyme reaction solution.
8. The reaction system of claim 7 , wherein the more than one glucosyltransferase enzyme comprises a mixture of at least one primer-dependent enzyme and at least one primer-independent enzyme.
9. The reaction system of claim 1 , wherein the poly (alpha 1,3 glucan) is a solid.
10. The reaction system of claim 1 , wherein the second chamber comprises a sucrose solution.