Methods for production of oligosaccharides
AssigneeZimitech
InventorsJingjing Liu, James Harrison Doudna Cate
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Abstract
Disclosed herein are genetically modified microorganisms and related methods for the enhanced production and export of oligosaccharides. The microorganisms described herein express major facility superfamily proteins such as CDT-1, which allows for the export of oligosaccharides. Variants of CDT-1 exhibit higher activity regarding oligosaccharide export. The microorganisms described herein express formation enzymes for the production of oligosaccharides. Means to export oligosaccharides into the growth medium are provided herein.
Claims68
An engineered microorganism capable of producing a human milk oligosaccharide (HMO) comprising:
- a first heterologous gene encoding an HMO formation enzyme and a second heterologous gene encoding a transporter for export of the HMO, wherein the transporter is CDT-1 or a variant thereof,
- wherein the HMO is a Lacto-N-Triose II (LNTII)-derived HMO or a sialylated HMO.
The engineered microorganism according to any of claim 1-12, wherein the engineered microorganism utilizes lactose as an HMO substrate.
An engineered microorganism capable of producing a human milk oligosaccharide (HMO) comprising:
- a first heterologous gene encoding an HMO formation enzyme and a second heterologous gene encoding a variant of CDT-1, wherein the CDT-1 variant comprises a sequence having one or more amino acid replacements at positions corresponding to amino acid positions 91, 209, 213, 256, 262, 335, 411 of SEQ ID NO:4, or the CDT-1 variant is selected from the group consisting of CDT-1 N209S F262Y, CDT-1 G91A, CDT-1 F213L, CDT-1 L256V, CDT-1 F335A, CDT-1 S411A, and CDT-1 N209S F262W, or the CDT-1 variant comprises an amino acid replacement at a position near the sugar substrate binding pocket and/or the PESPR motif (SEQ ID NO:
- 43), such as G336, Q337, N341, or G471;
- and
- wherein the engineered microorganism produces an HMO and is improved in the uptake of lactose into the microorganism as compared to a parent microorganism that lacks CDT-1, or a variant thereof.
Dependencies are inferred from the claim text.