Provided herein are methods of making microorganisms modified for increased xylose consumption as compared to unmodified microorganisms. The methods include providing xylose-consuming microorganisms comprising two or more copies of a nucleic acid sequence encoding xylose isomerase and two or more copies of a nucleic acid sequence encoding a xylose kinase, culturing the microorganisms in medium containing xylose and harvesting a portion of the microorganisms. These steps are repeated multiple times. The microorganisms are then isolated. The isolated microorganisms have increased xylose consumption rates compared to control xylose-consuming microorganisms. Also provided are a population of microorganisms made by the provided methods. Methods of culturing the population of microorganisms and methods of reducing xylitol production in cultures comprising the population of microorganisms are provided.

The invention relates to a method for preparing a yeast which is capable of simultaneously fermenting a pentose and a hexose sugar, the method comprising providing a yeast which comprises: one or more heterologous genes encoding an enzyme of a pentose metabolic pathway, a disruption of a gene encoding a ribulose-phosphate 3-epimerase and a disruption of a gene encoding a glucose-6-phosphate isomerase, and one or more overexpressed endogenous genes encoding an enzyme of the pentose phosphate pathway; and subjecting the yeast to evolutionary engineering on a medium comprising a hexose sugar and at least one pentose sugar, selecting for a yeast with improved growth rate when grown on a media comprising a hexose and at least one pentose sugar, so as to obtain an evolved yeast.

The present specification relates to a transformed yeast strain capable of simultaneously utilizing xylose and glucose as carbon sources, a preparation method thereof and a biofuel production method using the same. The transformed yeast strain transforms a wild-type yeast strain incapable of using xylose as a carbon source and simultaneously convert glucose and xylose, thereby enabling high yield production of a biofuel. The economics and sustainability of the biofuel and biomaterial production processes can be highly enhanced by providing a strain which can easily be converted to a strain capable of producing a biofuel/material in a high yield through an additional modification.

The present invention relates to the provision of an enzymatic method for the preparation of 2,6-bis(hydroxymethyl) pyridine (Formula I) using as substrate 2,6-Dimethlypyridine (2,6-lutidene) and the multicomponent xylene monooxygenase comprising XylM and XylA from Pseudomonas putida (Arthrobacter siderocapsulatus). The enzymatic method of the present invention is advantageous over conventional synthetic preparations, providing access to the title compound with a one-step enzymatic procedure.

Acetate is a potent microbial inhibitor which can affect the performance of yeast in ethanolic fermentation. The present disclosure provides a recombinant microbial host cell having (i) a first genetic modification for increasing the activity of one or more proteins that function in a first metabolic pathway to convert acetate into an alcohol in the microbial host cell; (ii) a second genetic modification for increasing the activity of one or more proteins that function in a second metabolic pathway to import glycerol in the recombinant microbial host cell (iii) a third genetic modification for increasing the activity of one or more proteins that function in a third metabolic pathway to convert a C5 carbohydrate into ethanol in the microbial host cell. The recombinant microbial host cell comprises and natively expresses native proteins that function in a fourth native metabolic pathway to produce glycerol in the microbial host cell.

Described herein are recombinant yeast cells expressing a xylulose kinase (XK) which are suitable for fermentation of pentoses. Also described are recombinant yeast cells with higher tolerance to formic and/or acetic acid and suitable for fermentation of pentoses. Also described are recombinant yeast cells expressing an enolase, a phosphofructokinase beta subunit, a 6-phosphofructo-2-kinase, a glucose-6-phosphate isomerase, a phosphoglycerate mutase and/or a triose-phosphate isomerase, and suitable for fermentation of pentoses. Also described are recombinant yeast cells expressing a a phosphoglucomutase and/or phosphoribomutase which are suitable for fermentation of pentoses. Further described are are methods of using or producing such recombinant yeast cells and related materials.

The present invention relates to processes for producing ethanol comprising saccharifying cellulosic or starch-containing material and fermenting the saccharified material with a fermenting microorganism to produce ethanol. The fermenting organism is Saccharomyces cerevisiae strain MBG5151 (deposited under Accession No. Y-67971 at the Agricultural Research Service Culture Collection (NRRL), Illinois 61604 U.S.A.), Saccharomyces cerevisiae strain MBG5248 (deposited under Accession No. Y-68015 at the Agricultural Research Service Culture Collection (NRRL), Illinois 61604 U.S.A.) or a fermenting organism that has properties that the same or about the same as that of Saccharomyces cerevisiae MBG5151 or MBG5248.

The present invention relates to a recombinant microorganism which is capable of simultaneously fermenting at least two sugars in a lignocellulosic saccharified liquid, and also capable of generating diol.

Acetate is a potent microbial inhibitor which can affect the performance of yeast in ethanolic fermentation. The present disclosure provides a recombinant microbial host cell having (i) a first genetic modification for increasing the activity of one or more proteins that function in a first metabolic pathway to convert acetate into an alcohol in the microbial host cell; (ii) a second genetic modification for increasing the activity of one or more proteins that function in a second metabolic pathway to import glycerol in the recombinant microbial host cell (iii) a third genetic modification for increasing the activity of one or more proteins that function in a third metabolic pathway to convert a C5 carbohydrate into ethanol in the microbial host cell. The recombinant microbial host cell comprises and natively expresses native proteins that function in a fourth native metabolic pathway to produce glycerol in the microbial host cell.

A method for preparing a yeast capable of simultaneously fermenting a pentose and a hexose sugar, comprising: (a) providing a yeast comprising: one or more heterologous genes encoding an enzyme of a pentose metabolic pathway; disruptions of a gene encoding a ribulose-phosphate 3-epimerase and of a gene encoding a glucose-6-phosphate isomerase; one or more overexpressed endogenous genes encoding an enzyme of the pentose phosphate pathway; and a disruption of one or more genes encoding an NADPH dependent 6-phosphogluconate dehydrogenase, (b) subjecting said yeast to evolutionary engineering on a medium comprising a hexose sugar and at least one pentose sugar, selecting for a yeast with improved growth rate obtain an evolved yeast; (d) restoring, in the evolved yeast, one or more of the disrupted genes, or: (d) identifying genetic permutations in at least part of the genome of the evolved yeast by genome sequencing; (e) constructing an improved pentose and hexose-fermenting yeast comprising one or more said genetic permutations. Also described is a recombinant yeast comprising one or more heterologous genes of a pentose metabolic pathway, and a gene encoding a variant of a parent polypeptide, the variant comprising an amino acid sequence comprising at least one mutation, when aligned with the amino acid sequence in SEQ ID NO: 6.