Anthocyanin bioproduction in a cell-free manufacturing system
AssigneeDebut Biotechnology, Inc.
InventorsJoshua Britton, Nicholas Brideau, Neill Kim
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Abstract
A cell-free system is provided to produce anthocyanins from anthocyanidins. An anthocyanidin is added to a reaction mixture comprising a glycosyl transferase enzyme and UDP-sugar. The reaction mixture may be aqueous and may comprise a co-solvent, e.g., an organic co-solvent, such as DMF.
Claims44
A method of producing an anthocyanin from an anthocyanidin, the method comprising:
- a) adding to a mixture comprising a suitable solvent (i) an anthocyanidin of Formula I:
- wherein each of R3, R5, R6, R7, R3′, R4′, and R5′ are independently selected from the group consisting of H, OH, and OCH3, wherein at least one of R3, R5, R6, R7, R3′, R4′, and R5′ is OH or CH3 and at least one of R3, R5, R6, R7, R3′, R4 ′, and R5′ is OH;
- (ii) a glycosylated uridine-5′-diphosphate of Formula II:
- wherein G is a glycosyl residue selected from the group consisting of 5- and 6-membered monosaccharides;
- and
- (iii) a glycosyltransferase enzyme to form a reaction mixture;
- b) removing the supernatant from the reaction mixture from (a);
- and
- c) isolating the anthocyanin.
A method of one of claims 1 to 14 having a starting pH, wherein the starting pH is about 4 to about 6.
A method of producing an anthocyanin from an anthocyanidin, comprising:
- a) in a cell-free vessel, providing a glycosyltransferase enzyme;
- b) adding an anthocyanidin and a glycosylated uridine diphosphate to the cell-free vessel to form the anthocyanin;
- and
- c) removing the anthocyanin from the cell-free vessel.
A method of producing an anthocyanin from an anthocyanidin, the method comprising:
- a) adding an anthocyanidin or anthocyanin of Formula Ia:
- wherein each of R3, R5, R6, R7, R3′, R4′, and R5′ are independently selected from the group consisting of H, OH, OCH3, or a sugar, wherein at least one of R3, R5, R6, R7, R3′, R4′, and R5′ is OH;
- a glycosylated uridine-5′-diphosphate of Formula II:
- wherein G is a glycosyl residue selected from the group consisting of 5- and 6-membered monosaccharides;
- and
- a glycosyltransferase enzyme to form a reaction mixture;
- b) removing the supernatant from the reaction mixture from (a);
- and
- c) isolating the anthocyanin.
5%, atleast95%, atleast 97.5%, atleast99%, atleast99.5%, or atleast99.9% similarity to SEQ ID NO 1-5.
Dependencies are inferred from the claim text.