Abstract: The present invention provides a method of identifying an enzyme, the method generally involving contacting a sample containing an enzyme with a selected enzyme substrate, where the contacting provides for covalent binding of the substrate to an amino acid of the enzyme to form a covalently modified enzyme; and determining the amino acid sequence of at least a portion of the covalently modified enzyme, using any available peptide sequencing technology, such as tandem mass spectrometry. The present invention further provides methods of identifying a nucleic acid encoding an enzyme, the methods generally involving identifying an enzyme; and, based on the amino acid sequence of at least a portion of the enzyme, designing nucleic acid probes or primers that hybridize to the nucleic acid encoding the enzyme.

Abstract: Methods for synthesizing isopentenyl pyrophosphate are provided. A first method comprises introducing into a host microorganism a plurality of heterologous nucleic acid sequences, each coding for a different enzyme in the mevalonate pathway for producing isopentenyl pyrophosphate. A related method comprises introducing into a host microorganism an intermediate in the mevalonate pathway and at least one heterologous nucleic acid sequence, each sequence coding for an enzyme in the mevalonate pathway necessary for converting the intermediate into isopentenyl pyrophosphate. The invention also provides nucleic acid sequences, enzymes, expression vectors, and transformed host cells for carrying out the methods.

Abstract: The present invention provides genetically modified host cells and use of same for producing isoprenoid compounds.

Abstract: Methods for synthesizing isopentenyl pyrophosphate are provided. A first method comprises introducing into a host microorganism a plurality of heterologous nucleic acid sequences, each coding for a different enzyme in the mevalonate pathway for producing isopentenyl pyrophosphate. A related method comprises introducing into a host microorganism an intermediate in the mevalonate pathway and at least one heterologous nucleic acid sequence, each sequence coding for an enzyme in the mevalonate pathway necessary for converting the intermediate into isopentenyl pyrophosphate. The invention also provides nucleic acid sequences, enzymes, expression vectors, and transformed host cells for carrying out the methods.

Abstract: The present invention provides methods of increasing production of an isoprenoid or an isoprenoid precursor in a host cell, the methods generally involving modulating the level of activity of a fatty acid biosynthetic pathway enzyme in the host cell and/or culturing the host cell in a culture medium comprising a fatty acid or a compound that can be metabolized in a cell or broken down in the medium to yield a fatty acid and/or culturing the host cell in a culture medium having increased osmolarity.

Abstract: The present invention provides methods of producing an isoprenoid or an isoprenoid precursor in a genetically modified host cell. The methods generally involve modulating the level of hydroxymethylglutaryl-CoA (HMG-CoA) in the cell, such that the level of HMG-CoA is not toxic to the cell and/or does not substantially inhibit cell growth, but is maintained at a level that provides for high-level production of mevalonate, IPP, and other downstream products of an isoprenoid or isoprenoid pathway, e.g., polyprenyl diphosphates and isoprenoid compounds. The present invention further provides genetically modified host cells that are suitable for use in a subject method. The present invention further provides recombinant nucleic acid constructs for use in generating a subject genetically modified host cell, including recombinant nucleic acid constructs comprising nucleotide sequences encoding one or more mevalonate pathway enzymes, and recombinant vectors (e.g., recombinant expression vectors) comprising same.

Abstract: Methods for synthesizing isopentenyl pyrophosphate are provided. A first method comprises introducing into a host microorganism a plurality of heterologous nucleic acid sequences, each coding for a different enzyme in the mevalonate pathway for producing isopentenyl pyrophosphate. A related method comprises introducing into a host microorganism an intermediate in the mevalonate pathway and at least one heterologous nucleic acid sequence, each sequence coding for an enzyme in the mevalonate pathway necessary for converting the intermediate into isopentenyl pyrophosphate. The invention also provides nucleic acid sequences, enzymes, expression vectors, and transformed host cells for carrying out the methods.

Abstract: Methods for synthesizing isopentenyl pyrophosphate are provided. A first method comprises introducing into a host microorganism a plurality of heterologous nucleic acid sequences, each coding for a different enzyme in the mevalonate pathway for producing isopentenyl pyrophosphate. A related method comprises introducing into a host microorganism an intermediate in the mevalonate pathway and at least one heterologous nucleic acid sequence, each sequence coding for an enzyme in the mevalonate pathway necessary for converting the intermediate into isopentenyl pyrophosphate. The invention also provides nucleic acid sequences, enzymes, expression vectors, and transformed host cells for carrying out the methods.

Abstract: Methods for synthesizing isopentenyl pyrophosphate are provided. A first method comprises introducing into a host microorganism a plurality of heterologous nucleic acid sequences, each coding for a different enzyme in the mevalonate pathway for producing isopentenyl pyrophosphate. A related method comprises introducing into a host microorganism an intermediate in the mevalonate pathway and at least one heterologous nucleic acid sequence, each sequence coding for an enzyme in the mevalonate pathway necessary for converting the intermediate into isopentenyl pyrophosphate. The invention also provides nucleic acid sequences, enzymes, expression vectors, and transformed host cells for carrying out the methods.

Abstract: The present invention provides methods of designing and generating polypeptide variants that have altered function compared to a parent polypeptide. The present invention further provides a computer program product for carrying out the design of a variant polypeptide. The present invention further provides nucleic acids encoding terpene cyclase variants, as well as vectors and host cells comprising the nucleic acids. The present invention further provides variant terpene cyclases; methods of producing the variant terpene cyclases; and methods of producing isoprenoid compounds.

Abstract: The present invention provides nucleic acids that include a promoter that is inducible by a transcriptional activator protein; and a nucleotide sequence that encodes the transcriptional activator protein. The present invention provides expression vectors that provide for inducible production of gene products in a host cell. The present invention further provides host cells genetically modified with a subject expression vector. The present invention further provides methods for producing a gene product in a host cell.

Abstract: The present invention provides nucleic acids comprising nucleotide sequences encoding modified cytochrome P450 enzymes; as well as recombinant vectors and host cells comprising the nucleic acids. The present invention further provides methods of producing a functionalized compound in a host cell genetically modified with a nucleic acid comprising nucleotide sequences encoding a modified cytochrome P450 enzyme.

Abstract: The present invention provides genetically modified eukaryotic host cells exhibiting increased activity levels of one or more enzymes that generate precursors to be utilized by the mevalonate pathway enzymes, increased activity levels of one or more mevalonate pathway enzymes, prenyl transferase, and/or decreased levels of squalene synthase activity; such cells are useful for producing isoprenoid compounds. The present invention provides genetically modified eukaryotic host cells that produce higher levels of acetyl-CoA than a control cell; such cells are useful for producing a variety of products, including isoprenoid compounds. Methods are provided for the production of an isoprenoid compound or an isoprenoid precursor in a subject genetically modified eukaryotic host cell. The methods generally involve culturing a subject genetically modified host cell under conditions that promote production of high levels of an isoprenoid or isoprenoid precursor compound.

Abstract: The present invention provides methods of producing an isoprenoid or an isoprenoid precursor in a genetically modified host cell. The methods generally involve modulating the level of hydroxymethylglutaryl-CoA (HMG-CoA) in the cell, such that the level of HMG-CoA is not toxic to the cell and/or does not substantially inhibit cell growth, but is maintained at a level that provides for high-level production of mevalonate, IPP, and other downstream products of an isoprenoid or isoprenoid pathway, e.g., polyprenyl diphosphates and isoprenoid compounds. The present invention further provides genetically modified host cells that are suitable for use in a subject method. The present invention further provides recombinant nucleic acid constructs for use in generating a subject genetically modified host cell, including recombinant nucleic acid constructs comprising nucleotide sequences encoding one or more mevalonate pathway enzymes, and recombinant vectors (e.g., recombinant expression vectors) comprising same.

Abstract: The present invention provides methods of designing and generating polypeptide variants that have altered properties compared to a parent polypeptide. The present invention further provides a computer program product for carrying out the design of a variant polypeptide. The present invention further provides nucleic acids encoding enzyme variants, as well as vectors and host cells comprising the nucleic acids. The present invention further provides variant enzymes; methods of producing the variant enzymes; and methods of producing compounds using the enzymes.

Abstract: The present invention provides genetically modified host cells that exhibit modified activity levels of one or more gene products such that, when a cytochrome P450 enzyme is produced in the genetically modified host cell, the modified activity levels of the one or more gene products provide for enhanced production and/or activity of the cytochrome P450 enzyme. The present invention provides methods of producing a cytochrome P450 enzyme in a host cell, generally involving culturing a subject genetically modified host cell in a suitable culture medium. The present invention further provides methods of producing a product of a P450-dependent oxidation, generally involving culturing a subject genetically modified host cell in a suitable culture medium.

Abstract: The present invention provides artemisinic epoxide, and methods of synthesizing artemisinic epoxide in a genetically modified host cell. The present invention further provides methods for producing artemisinin. The present invention further provides variant enzymes that catalyze the oxidation of amorpha-4,11-diene to artemisinic epoxide; nucleic acids encoding the variant enzymes; as well as recombinant vectors and host cells comprising the nucleic acids.

Abstract: The present invention provides genetically modified eukaryotic host cells that produce isoprenoid precursors or isoprenoid compounds. A subject genetically modified host cell comprises increased activity levels of one or more of mevalonate pathway enzymes, increased levels of prenyltransferase activity, and decreased levels of squalene synthase activity. Methods are provided for the production of an isoprenoid compound or an isoprenoid precursor in a subject genetically modified eukaryotic host cell. The methods generally involve culturing a subject genetically modified host cell under conditions that promote production of high levels of an isoprenoid or isoprenoid precursor compound.

Abstract: The present invention provides methods of producing an isoprenoid or an isoprenoid precursor in a genetically modified host cell. The methods generally involve modulating the level of hydroxymethylglutaryl-CoA (HMG-CoA) in the cell, such that the level of HMG-CoA is not toxic to the cell and/or does not substantially inhibit cell growth, but is maintained at a level that provides for high-level production of mevalonate, IPP, and other downstream products of an isoprenoid or isoprenoid pathway, e.g., polyprenyl diphosphates and isoprenoid compounds. The present invention further provides genetically modified host cells that are suitable for use in a subject method. The present invention further provides recombinant nucleic acid constructs for use in generating a subject genetically modified host cell, including recombinant nucleic acid constructs comprising nucleotide sequences encoding one or more mevalonate pathway enzymes, and recombinant vectors (e.g., recombinant expression vectors) comprising same.

Abstract: The invention provides methods and microbial cultures for the bioremediation of organic halide contaminated groundwater contaminated with organic halides, such as di- and trichloroethene. The methods involve adding, in situ to organic halide-contaminated groundwater a carbohydrate and one or more reductive dehalogenation factors, usually in the form of a nutrient extract, both in amounts sufficient to permit in situ reductive dehalogenation of the organic halide by a microbial population. The microbial population may be endogenous to the ground water or added exogenously. The nutrient-enriched ground water is then maintained in situ under reducing conditions to reductively dehalogenate the contaminating organic halide. Enriched bioremediation cultures are produced by adding to organic halide contaminated groundwater which comprises an endogenous microbial population capable of reductive dehalogenation of the organic halide a carbohydrate and frequently, one or more reductive dehalogenation factors.