Glycosyltransferase glycosylating flavokermesic acid and/or kermesic acid

Patent metadata

Field	Value
Publication number	US-10724012-B2
Application number	US-201816160396-A
Country	US
Kind code	B2
Filing date	Oct 15, 2018
Priority date	Dec 18, 2013
Publication date	Jul 28, 2020
Grant date	Jul 28, 2020

Abstract

An isolated glycosyltransferase (GT) polypeptide capable of: (I): conjugating glucose to flavokermesic acid (FK); and/or (II): conjugating glucose to kermesic acid (KA) and use of this GT to e.g. make Carminic acid.

First claim

We claim:

1. A method for producing flavokermesic acid glycoside or kermesic acid glycoside comprising the acts of:
providing an aglycon wherein said aglycon comprises flavokermesic acid or kermesic acid;
providing a sugar, wherein said sugar comprises a nucleotide activated glucose;
providing a recombinant cell wherein said recombinant cell comprises an expression cassette comprising an isolated polynucleotide comprising a nucleotide sequence which encodes an isolated glycosyltransferase polypeptide capable of:
(I): conjugating nucleotide activated glucose to flavokermesic acid (FK); and/or
(II): conjugating nucleotide activated glucose to kermesic acid (KA);

and wherein the glycosyltransferase polypeptide is at least one polypeptide selected from the group consisting of:
(a) a polypeptide comprising an amino acid sequence which has at least 95% identity with amino acids 1 to 515 of SEQ ID NO:2; and
(b) a polypeptide comprising an amino acid sequence which has at least 95% identity with amino acids 20 to 468 of SEQ ID NO:2; and

wherein said polynucleotide is operably linked to one or more control sequences that direct the production of the polypeptide in an expression host; and
contacting said glycosyltransferase polypeptide with said aglycon and said nucleotide activated glucose to produce flavorkermesic acid glycoside or kermesic acid glycoside.

2. The method of claim 1 , wherein said cell is Saccharomyces spp. or Pichia spp.

3. The method of claim 2 , wherein said cell is Saccharomyces cerevisiae.

4. The method of claim 1 , wherein said nucleotide activated glucose comprises UDP-glucose.

5. The method of claim 1 , wherein said expression cassette comprises a plasmid.

6. The method of claim 1 , further comprising the act of purifying said produced flavokermesic acid glycoside or kermesic acid glycoside.

7. A method for producing flavokermesic acid glycoside or kermesic acid glycoside in vitro comprising the acts of:
providing an aglycon wherein said aglycon comprises flavokermesic acid or kermesic acid;
providing a sugar, wherein said sugar comprises a nucleotide activated glucose;
providing an isolated recombinant glycosyltransferase produced by recombinant cell wherein said recombinant cell comprises an expression cassette comprising an isolated polynucleotide comprising a nucleotide sequence which encodes an isolated glycosyltransferase polypeptide capable of:
(I): conjugating nucleotide activated glucose to flavokermesic acid (FK); and/or
(II): conjugating nucleotide activated glucose to kermesic acid (KA);

and wherein the glycosyltransferase polypeptide is at least one polypeptide selected from the group consisting of:
(a) a polypeptide comprising an amino acid sequence which has at least 95% identity with amino acids 1 to 515 of SEQ ID NO:2; and
(b) a polypeptide comprising an amino acid sequence which has at least 95% identity with amino acids 20 to 468 of SEQ ID NO:2; and

wherein said polynucleotide is operably linked to one or more control sequences that direct the production of the polypeptide in an expression host; and
contacting said glycosyltransferase polypeptide with said aglycon and said nucleotide activated glucose to produce flavokermesic acid glycoside or kermesic acid glycoside.

8. The method of claim 7 , wherein said cell is Saccharomyces spp. or Pichia spp.

9. The method of claim 8 , wherein said cell is Saccharomyces cerevisiae.

10. The method of claim 7 , wherein said nucleotide activated glucose comprises UDP-glucose.

11. The method of claim 7 , wherein said isolated recombinant glycosyltransferase is isolated by the act of extraction from said recombinant cell comprising a expression cassette which encodes for a glycosyltransferase.

Assignees

• Univ Copenhagen
• Univ Danmarks Tekniske

Inventors

• Kannangara Rubini Maya
• Bennedsen Mads
• Madsen Bjørn
• Binderup Kim
• Thrane Ulf
• Frandsen Rasmus John Normand
• Mortensen Uffe Hasbro
• Møller Birger Lindberg
• Okkels Finn Thyge

Classifications

• C12P7/66

  containing the quinoid structure · CPC title

• C12N9/1051

  Hexosyltransferases (2.4.1) · CPC title

• C12Y204/00

  Glycosyltransferases (2.4) · CPC title

• C12N9/1048 Primary

  Glycosyltransferases (2.4) · CPC title

• C12N15/81

  for yeasts · CPC title