Oligosaccharide production in yeast
AssigneeAmyris, Inc.
InventorsJessica WALTER, Dominic PINEL
Life of this patent
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Abstract
Provided herein are genetically modified yeast cells capable of producing one or more human milk oligosaccharides (HMOs) and methods of making such cells. The yeast cells are engineered to comprise a heterologous nucleic acid encoding a transporter protein and one or more heterologous nucleic acids that encode enzymes of a HMO biosynthetic pathway. Also provided are fermentation compositions including the disclosed genetically modified yeast cells, and related methods of producing and recovering HMOs generated by the yeast cells.
Claims21
1-99. (canceled)
The yeast cell of claim 100, wherein the ABC transporter has at least 95% identity to any one of SEQ ID NOS:
- 28-98.
The yeast cell of claim 100, wherein the ABC transporter has at least 95% identity to any one of SEQ ID NOS:
- 28-55.
The yeast cell of claim 100, wherein the ABC transporter has at least 95% identity to any one of SEQ ID NOS:
- 28-38 and 55.
The yeast cell of claim 100, wherein the ABC transporter has at least 95% identity to any one of SEQ ID NO:
- 28, SEQ ID NO:
- 29, SEQ ID NO:
- 30, SEQ ID NO:
- 31, SEQ ID NO:
- 32, and SEQ ID NO:
- 55.
A yeast cell genetically modified to produce one or more human milk oligosaccharides, wherein the yeast cell comprises (i) a heterologous nucleic acid encoding a human milk oligosaccharide (HMO) ABC transporter polypeptide;
- and (ii) one or more heterologous nucleic acids that each independently encode at least one enzyme of a human milk oligosaccharide biosynthetic pathway, wherein the ABC transporter exports the human milk oligosaccharide 6′-siallylactose.
The yeast cell of claim 100, wherein the one or more human milk oligosaccharides comprise 2′-fucosyllactose, 3-fucosyllactose, lacto-N-tetraose, lacto-N-neotetraose, 3′-sialyllactose, 6′-sialyllactose, and/or difucosyllactose.
The yeast cell of claim 100, wherein the yeast cell is a Saccharomyces sp. or a Kluveromyces sp.
The yeast cell of claim 100, wherein the enzymes encoded by the one or more heterologous nucleic acids further comprise a protein that transports lactose into the yeast cell.
A method of producing one or more human milk oligosaccharides, the method comprising culturing a population of genetically modified yeast cells of claim 100 in a culture medium under conditions suitable for the yeast cells to produce the one or more human milk oligosaccharides.
A fermentation composition comprising:
- a population of genetically modified yeast cells comprising the yeast cell of claim 100;
- and a culture medium comprising one or more human milk oligosaccharides produced from the yeast cells.
A method of genetically modifying a yeast cell to produce one or more human milk oligosaccharides, the method comprising:
- (a) (i) introducing a heterologous nucleic acid encoding an ABC transporter polypeptide into the yeast cell, wherein the ABC transporter polypeptide exports the human milk oligosaccharide lacto-N-neotetraose;
- and (ii) introducing one or more heterologous nucleic acids that each independently encode at least one enzyme of a human milk oligosaccharide biosynthetic pathway into the yeast cell;
- or
- (b) introducing a heterologous nucleic acid encoding an ABC transporter polypeptide into the yeast cell, wherein the ABC transporter polypeptide exports the human milk oligosaccharide lacto-N-neotetraose, and wherein the yeast cell comprises one or more heterologous nucleic acids that each independently encode at least one enzyme of a human milk oligosaccharide biosynthetic pathway into the yeast cell.
A yeast cell genetically modified to produce one or more human milk oligosaccharides, wherein the yeast cell comprises (i) a heterologous nucleic acid encoding a human milk oligosaccharide (HMO) ABC transporter polypeptide;
- and (ii) one or more heterologous nucleic acids that each independently encode at least one enzyme of a human milk oligosaccharide biosynthetic pathway, wherein the ABC transporter exports the human milk oligosaccharide lacto-N-neotetraose.
Dependencies are inferred from the claim text.