Abstract: The present invention provides for heterologous expression of termite and termite-associated symbiont cellulases. The cellulases can, for example, be codon-optimized and expressed in yeast host cells, such as the yeast Saccharomyces cerevisiae. The cellulases can also be co-expressed in host cells with other cellulases. The expression in such host cells of the termite and termite-associated symbiont cellulases, and variants and combinations thereof, result in yeast with improved cellulosic activity. Thus, such genes and expression systems are useful for efficient and cost-effective consolidated bioprocessing systems.

Abstract: One aspect of the invention relates to a genetically modified thermophilic or mesophilic microorganism, wherein a first native gene is partially, substantially, or completely deleted, silenced, inactivated, or down-regulated, which first native gene encodes a first native enzyme involved in the metabolic production of an organic acid or a salt thereof, thereby increasing the native ability of said thermophilic or mesophilic microorganism to produce ethanol as a fermentation product. In certain embodiments, the aforementioned microorganism further comprises a first non-native gene, which first non-native gene encodes a first non-native enzyme involved in the metabolic production of ethanol. Another aspect of the invention relates to a process for converting lignocellulosic biomass to ethanol, comprising contacting lignocellulosic biomass with a genetically modified thermophilic or mesophilic microorganism.

Abstract: One aspect of the invention relates to industrial bioconversion of the xylose portion of biomass materials into fuels and chemicals. Another aspect of the invention relates to industrial bioconversion of the xylan portion of biomass materials into fuels and chemicals. In one embodiment, the invention is directed to the bacterium Clostridium thermocellum, a highly cellulolytic organism that has much potential as a biocatalyst in a consolidated bioprocess configuration. In some embodiments, the invention is a genetic modification that confers the ability to ferment xylose to C. thermocellum and the strains created with this modification. In some embodiments, the genetic modification is composed of two genes contained in an operon from T. saccharolyticum. The genes express proteins with xylose isomerase (XI) and xylulokinase activites (XK). In other embodiments, the invention relates to a recombinant Clostridium thermocellum host cell capable of fermenting xylan.

Abstract: The present invention provides for the manipulation of carbon flux in a recombinant host cell to increase the formation of desirable products. The invention relates to cellulose-digesting organisms that have been genetically modified to allow the production of ethanol at a high yield by redirecting carbon flux at key steps of central metabolism.

Abstract: The present invention is directed to a yeast strain, or strains, secreting a full suite, or any subset of that full suite, of enzymes to hydrolyze corn starch, corn fiber, lignocellulose, (including enzymes that hydrolyze linkages in cellulose, hemicellulose, and between lignin and carbohydrates) and to utilize pentose sugars (xylose and arabinose). The invention is also directed to the set of proteins that are well expressed in yeast for each category of enzymatic activity. The resulting strain, or strains can be used to hydrolyze starch and cellulose simultaneously. The resulting strain, or strains can be also metabolically engineered to produce less glycerol and uptake acetate. The resulting strain, or strains can also be used to produce ethanol from granular starch without liquefaction.

Abstract: The present invention provides for novel metabolic pathways to reduce or eliminate glycerol production and increase product formation. More specifically, the invention provides for a recombinant microorganism comprising a deletion of one or more native enzymes that function to produce glycerol and/or regulate glycerol synthesis and one or more native and/or heterologous enzymes that function in one or more engineered metabolic pathways to convert a carbohydrate source, such as lignocellulose, to a product, such as ethanol, wherein the one or more native and/or heterologous enzymes is activated, upregulated, or downregulated.

Abstract: The present invention is directed to a process for biologically converting carbohydrates from lignocellulosic biomass comprising the steps of: suspending lignocellulosic biomass in a flow-through reactor, passing a reaction solution into the reactor, wherein the solution is absorbed into the biomass substrate and at least a portion of the solution migrates through said biomass substrate to a liquid reservoir, recirculating the reaction solution in the liquid reservoir at least once to be absorbed into and migrate through the biomass substrate again. The biological converting of the may involve hydrolyzing cellulose, hemicellulose, or a combination thereof to form oligosaccharides, monomelic sugars, or a combination thereof; fermenting oligosaccharides, monomelic sugars, or a combination thereof to produce ethanol, or a combination thereof. The process can further comprise removing the reaction solution and processing the solution to separate the ethanol produced from non-fermented solids.

Abstract: The present invention provides for heterologous expression of termite and termite-associated symbiont cellulases. The cellulases can, for example, be codon-optimized and expressed in yeast host cells, such as the yeast Saccharomyces cerevisiae. The cellulases can also be co-expressed in host cells with other cellulases. The expression in such host cells of the termite and termite-associated symbiont cellulases, and variants and combinations thereof, result in yeast with improved cellulosic activity. Thus, such genes and expression systems are useful for efficient and cost-effective consolidated bioprocessing systems.

Abstract: The present invention is directed to a yeast strain, or strains, secreting a full suite, or any subset of that full suite, of enzymes to hydrolyze corn starch, corn fiber, lignocellulose, (including enzymes that hydrolyze linkages in cellulose, hemicellulose, and between lignin and carbohydrates) and to utilize pentose sugars (xylose and arabinose). The invention is also directed to the set of proteins that are well expressed in yeast for each category of enzymatic activity. The resulting strain, or strains can be used to hydrolyze starch and cellulose simultaneously. The resulting strain, or strains can be also metabolically engineered to produce less glycerol and uptake acetate. The resulting strain, or strains can also be used to produce ethanol from granular starch without liquefaction.

Abstract: The present invention provides for novel metabolic pathways to convert biomass and other carbohydrate sources to malonyl-CoA derived products, such as hydrocarbons and other bioproducts, under anaerobic conditions and with the net production of ATP. More specifically, the invention provides for a recombinant microorganism comprising one or more native and/or heterologous enzymes that function in one or more engineered metabolic pathways to achieve conversion of a carbohydrate source to, e.g., long-chain hydrocarbons and hydrocarbon derivatives, wherein the one or more native and/or heterologous enzymes is activated, upregulated, downregulated, or deleted. The invention also provides for processes to convert biomass to malonyl-CoA derived products which comprise contacting a carbohydrate source with a recombinant microorganism of the invention.

Abstract: One aspect of the invention relates to a genetically modified thermophilic or mesophilic microorganism, wherein a first native gene is partially, substantially, or completely deleted, silenced, inactivated, or down-regulated, which first native gene encodes a first native enzyme involved in the metabolic production of an organic acid or a salt thereof, thereby increasing the native ability of said thermophilic or mesophilic microorganism to produce lactate or acetate as a fermentation product. In certain embodiments, the aforementioned microorganism further comprises a first non-native gene, which first non-native gene encodes a first non-native enzyme involved in the metabolic production of lactate or acetate. Another aspect of the invention relates to a process for converting lignocellulosic biomass to lactate or acetate, comprising contacting lignocellulosic biomass with a genetically modified thermophilic or mesophilic microorganism.

Abstract: The present invention provides for the isolation and characterization of the cbh1 gene from Schizochytrium aggregatum. In particular, the present invention provides for the nucleic acid and amino acid sequences of Schizochytrium aggregatum cbh1, and domains, variants and derivatives thereof. The present invention further provides for the heterologous expression of Schizochytrium aggregatum Cbh1 in host cells, including yeast, e.g., Saccharomyces cerevisiae. Expression of Schizochytrium aggregatum Cbh1 in host cells will augment cellulose digestion and facilitate ethanol production by those host cells on cellulosic substrates. In certain embodiments, heterologous expression in Saccharomyces cerevisiae is in coordination with heterologous expression of other known, or newly identified saccharolytic enzymes. Therefore, the present invention also provides that the novel Schizochytrium aggregatum Cbh1 gene can utilized in a consolidated bioprocessing system.

Abstract: One aspect of the invention relates to a genetically modified thermophilic or mesophilic microorganism, wherein a first native gene is partially, substantially, or completely deleted, silenced, inactivated, or down-regulated, which first native gene encodes a first native enzyme involved in the metabolic production of an organic acid or a salt thereof, thereby increasing the native ability of said thermophilic or mesophilic microorganism to produce ethanol as a fermentation product. In certain embodiments, the aforementioned microorganism further comprises a first non-native gene, which first non-native gene encodes a first non-native enzyme involved in the metabolic production of ethanol. Another aspect of the invention relates to a process for converting lignocellulosic biomass to ethanol, comprising contacting lignocellulosic biomass with a genetically modified thermophilic or mesophilic microorganism.

Abstract: The present invention provides for novel metabolic pathways leading to acrylate formation in a consolidated bio-processing system (CBP) where lignocellulosic biomass is efficiently converted to acrylate. In one such metabolic pathway, pyruvate is converted to lactate, which is converted to lactoyol-CoA, which is converted to acryloyl-CoA, and which is finally converted to acrylate. In another such metabolic pathway, pyruvate is converted to L-?-alanine, which is converted to L-aspartate, which is converted to ?-alanine, which is converted to ?-alanyl-CoA, which is converted to acryloyl-CoA, and which is finally converted to acrylate. In yet another metabolic pathway, pyruvate is converted to lactate, and then lactate is converted directly to acrylate.

Abstract: The present invention is directed to a process of producing substantially pure lignin from lignocellulosic biomass, which comprises: pre-treating a lignocellulosic feedstock to produce a reactive lignin-carbohydrate mixture; biologically-reacting the carbohydrates in the mixture, separating remaining solids from the liquid fermentation products, and drying the resulting solids to yield a substantially pure lignin product. Optionally, the lignin product may be washed and subjected to a second hydrolysis step. Optionally, the lignin product may be further processed by hydrotreating and/or pyrolysis in order to yield desirable products such as fuel additives.

Abstract: One aspect of the invention relates to a genetically modified thermophilic or mesophilic microorganism, wherein a first native gene is partially, substantially, or completely deleted, silenced, inactivated, or down-regulated, which first native gene encodes a first native enzyme involved in the metabolic production of an organic acid or a salt thereof, thereby increasing the native ability of said thermophilic or mesophilic microorganism to produce lactate or acetate as a fermentation product. In certain embodiments, the aforementioned microorganism further comprises a first non-native gene, which first non-native gene encodes a first non-native enzyme involved in the metabolic production of lactate or acetate. Another aspect of the invention relates to a process for converting lignocellulosic biomass to lactate or acetate, comprising contacting lignocellulosic biomass with a genetically modified thermophilic or mesophilic microorganism.

Abstract: Thermophilic gram-positive anaerobic host cells, for example Thermoanaerobacterium saccharolyticum (“T sacch”), express heterologous biomass degrading enzymes, such as cellulases, and are able to produce useful fermentation products from cellulose. Useful fermentation products include, for example, ethanol, acetic acid, lactic acid or CO2. In order to provide maximum expression and activity levels, biomass degrading enzymes can be expressed from codon-optimized nucleotide sequences, can be expressed under the control of a high-efficiency promoter, and/or can be fused to a signal peptide. In addition, the host cell, for example, a T sacch host cell, can be genetically altered to further improve ethanol production, for example by disrupting the production of organic products other than ethanol.

Abstract: The present invention provides for heterologous expression of termite and termite-associated symbiont cellulases. The cellulases can, for example, be codon-optimized and expressed in yeast host cells, such as the yeast Saccharomyces cerevisiae. The cellulases can also be co-expressed in host cells with other cellulases. The expression in such host cells of the termite and termite-associated symbiont cellulases, and variants and combinations thereof, result in yeast with improved cellulosic activity. Thus, such genes and expression systems are useful for efficient and cost-effective consolidated bioprocessing systems.

Abstract: The present invention provides for the isolation and characterization of the cbh1 gene from Schizochytrium aggregatum. In particular, the present invention provides for the nucleic acid and amino acid sequences of Schizochytrium aggregatum cbh1, and domains, variants and derivatives thereof. The present invention further provides for the heterologous expression of Schizochytrium aggregatum Cbh1 in host cells, including yeast, e.g., Saccharomyces cerevisiae. Expression of Schizochytrium aggregatum Cbh1 in host cells will augment cellulose digestion and facilitate ethanol production by those host cells on cellulosic substrates. In certain embodiments, heterologous expression in Saccharomyces cerevisiae is in coordination with heterologous expression of other known, or newly identified saccharolytic enzymes. Therefore, the present invention also provides that the novel Schizochytrium aggregatum Cbh1 gene can utilized in a consolidated bioprocessing system.

Abstract: The present invention relates to methods for transforming Gram-positive, anaerobic, thermophilic bacteria via electroporation and Gram-positive, anaerobic, thermophilic bacteria transformed by the disclosed methods. The methods employ voltage pulsing schemes that decrease arcing such that increased transformation efficiency and cell viability is observed. The present invention is further directed to a method for transforming Gram-positive, anaerobic, thermophilic bacteria via electroporation using recovery/selection temperatures to effect increased transformation efficiency in difficult to transform bacteria.