Limiting yeast-produced trehalose in fermentation

Patent metadata

Field	Value
Publication number	US-10947568-B2
Application number	US-202016776389-A
Country	US
Kind code	B2
Filing date	Jan 29, 2020
Priority date	Nov 6, 2015
Publication date	Mar 16, 2021
Grant date	Mar 16, 2021

Abstract

The present disclosure relates to recombinant yeast host cells having (i) a first genetic modification for reducing the production of one or more native enzymes that function to produce glycerol or regulating glycerol synthesis and/or allowing the production of an heterologous glucoamylase and (ii) a second genetic modification for reducing the production of one or more native enzymes that function to produce trehalose or regulating trehalose synthesis and/or allowing the expression of an heterologous trehalase. The recombinant yeast host cells can be used to limit the production of (yeast-produced) trehalose (particularly extracellular trehalose) during fermentation and, in some embodiments, can increase the production of a fermentation product (such as, for example, ethanol).

First claim

What is claimed is:

1. A process for converting a substrate into a fermentation product, the process comprising contacting biomass comprising the substrate with a recombinant yeast host cell under conditions to allow conversion of at least a part of the substrate into the fermentation product, wherein the recombinant yeast host cell comprises:
(i) a first genetic modification for reducing production of one or more native enzymes that function to produce glycerol or regulating glycerol synthesis and/or allowing production of an heterologous glucoamylase, wherein
the first genetic modification for reducing the production of the one or more native enzymes that function to produce glycerol and the one or more native enzymes that function to produce glycerol is glycerol-3-phosphate dehydrogenase 1 (GPD1), a polypeptide encoded by a gpd1 ortholog, glycerol-3-phosphate dehydrogenase 2 (GPD2) polypeptide and/or a polypeptide encoded by a gpd2 ortholog;
the first genetic modification for regulating glycerol synthesis is for reducing production of FDP1 suppressor (FPS1) polypeptide or a polypeptide encoded by a fps1 ortholog and/or is for increasing production of sugar transporter-like (STL1) polypeptide and/or a polypeptide encoded by a stl1 ortholog, and

(ii) a second genetic modification for reducing production of one or more native enzymes that function to produce trehalose or regulating trehalose synthesis and/or allowing expression of an heterologous trehalase, wherein
the second genetic modification for reducing the production of one or more native enzymes that function to produce trehalose and the one or more native enzymes that function to produce trehalose is trehalose-6-phosphate synthase 1 (TPS1), a polypeptide encoded by a tps1 ortholog, trehalose-6-phosphate synthase 2 (TPS2) polypeptide and/or a polypeptide encoded by a tps2 ortholog;
the second genetic modification for regulating trehalose synthesis is for reducing production of one or more native enzymes regulating trehalose synthesis and the one or more native enzymes regulating trehalose synthesis is trehalose phosphate synthase (TPS3), a polypeptide encoded by a tps3 ortholog, trehalose synthase long chain (TSL1) polypeptide, a polypeptide encoded by a tsl1 ortholog, neutral trehalase 1 (NTH1), a polypeptide encoded by a nth1 ortholog, neutral trehalase 2 (NTH2), a polypeptide encoded by a nth2 ortholog, acid trehalase 1 (ATH1), a polypeptide encoded by a ath1 ortholog, hexokinase 1 (HXK1), a polypeptide encoded by a hxk1 ortholog, hexokinase 2 (HXK2), a polypeptide encoded by a hxk2 ortholog, glucokinase 1 (GLK1), a polypeptide encoded by a glk1 ortholog, phosphoglucomutase 1 (PGM1), a polypeptide encoded by a pgm1 ortholog, phosphoglucomutase 2 (PGM2), a polypeptide encoded by a pgm2 ortholog, glycogen phosphorylase 1 (GPH1), a polypeptide encoded by a gph1 ortholog, UDP-glucose pyrophosphorylase (UGP1), a polypeptide encoded by a ugp1 ortholog, glycogen synthase 1 (GSY1), a polypeptide encoded by a gsy1 ortholog, glycogen synthase 2 (GSY2) and/or a polypeptide encoded by a gsy2 ortholog.

2. The process of claim 1 , wherein the recombinant yeast host cell has the first genetic modification for reducing the production of the one or more native enzymes that function to produce glycerol and the second genetic modification for reducing the production of the one or more native enzymes that function to produce trehalose or regulating trehalose synthesis.

3. The process of claim 1 , wherein the recombinant yeast host cell has the first modification for reducing the production of the one or more native enzymes that function to produce glycerol or regulating glycerol synthesis and the second modification allowing the production of the heterologous trehalase.

4. The process of claim 1 , wherein the recombinant yeast host cell has the first genetic modification allowing the production of the heterologous glucoamylase and the second genetic modification for reducing the production of the one or more native enzymes that function to produce trehalose or regulating trehalose synthesis.

5. The process of claim 1 , wherein the recombinant yeast host cell has the first genetic modification allowing the production of the heterologous glucoamylase and the second genetic modification allowing the production of the heterologous trehalase.

6. The process of claim 1 , wherein the recombinant yeast host cell further comprises a third genetic modification for reducing production of the one or more native enzymes that function to catabolize formate.

7. The process of claim 6 , wherein the recombinant yeast host cell lacks ability to produce a formate dehydrogenase 1 (FDH1) polypeptide and a formate dehydrogenase 2 (FDH2) polypeptide.

8. The process of claim 1 , wherein the recombinant yeast host cell further comprises a fourth genetic modification for reducing production of the one or more native enzymes that function to produce glycerol.

9. The process of claim 1 , wherein the recombinant yeast host cell further comprises a fifth genetic modification for regulating glycerol synthesis.

10. The process of claim 9 , wherein the recombinant yeast host cell has an ability to exhibit increased expression of a sugar transporter-like (STL1) polypeptide.

11. The process of claim 8 , wherein the heterologous glucoamylase has the amino acid sequence of SEQ ID NO: 3, is a variant of the amino acid sequence of SEQ ID NO: 3 having glucoamylase activity or is a fragment of the amino acid sequence of SEQ ID NO: 3 having glucoamylase activity.

12. The process of claim 1 , wherein the heterologous trehalase has the amino acid sequence of SEQ ID NO: 1 or 2, is a variant of the amino acid sequence of SEQ ID NO: 1 or 2 having acid trehalase activity or is a fragment of the amino acid sequence of SEQ ID NO: 1 or 2 having acid trehalase activity.

13. The process of claim 1 , wherein the recombinant yeast host cell is a cell from genus Saccharomyces sp.

14. The process of claim 11 , wherein the recombinant yeast host cell is a cell of species Saccharomyces cerevisiae.

15. The process of claim 1 , wherein the substrate comprises corn.

16. The process of claim 15 , wherein the corn is provided in a gelatinized form.

17. The process of claim 15 , wherein the corn is provided in a raw form.

18. The process of claim 13 , wherein the fermentation product is ethanol.

19. The process of claim 1 , wherein the substrate comprises maltodextrin.

20. The process of claim 1 , wherein the substrate comprises starch.

Assignees

• Lallemand Hungary Liquidity Man Llc

Inventors

• Rice Charles F
• Skinner Ryan
• Barrett Trisha
• Argyros Aaron

Classifications

• Y02E50/10

  Biofuels, e.g. bio-diesel · CPC title

• C12Y302/01028

  Alpha,alpha-trehalase (3.2.1.28) · CPC title

• C12P7/20

  Glycerol · CPC title

• C12N1/18

  Baker's yeast; Brewer's yeast · CPC title

• C12N15/81

  for yeasts · CPC title