P450 enzymes, nucleic acids encoding them and methods of making and using them
AssigneeDiversa Corporation
InventorsDavid Weiner, Mark J. Burk, Tim Hitchman, Catherine Pujol, Toby Richardson, Jay M. Short
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Abstract
The invention is directed to polypeptides having P450 activity, polynucleotides encoding the polypeptides, antibodies that bind to these polypeptides, and methods for making and using these polynucleotides and polypeptides. The P450 enzymes can be used to catalyze the hydrolysis of epoxides and arene oxides to their corresponding diols.
Claims55
An isolated, synthetic or recombinant nucleic acid, wherein the nucleic acid comprises
- (a) a sequence that hybridizes under stringent conditions to the complement of the polynucleotide comprising the nucleic acid sequence of SEQ ID NO:23, wherein the nucleic acid encodes a polypeptide having cytochrome P450 activity, and the stringent conditions comprise a wash step comprising a wash in 0.2×SSC at a temperature of about 65° C. for about 15 minutes, or
- (b) a sequence which is the complete complement of the polynucleotide of (a).
An isolated, synthetic or recombinant nucleic acid encoding a fusion protein comprising:
- (a) an epoxidase and an N-terminal or a carboxy-terminal heterologous peptide,
- wherein the epoxidase comprises a sequence having at least 95% sequence identity to SEQ ID NO:24, and wherein the fusion protein continues to have epoxidase activity.
An isolated, synthetic or recombinant nucleic acid comprising a nucleic acid sequence having at least 95% sequence identity to SEQ ID NO:23, wherein the nucleic acid encodes a polypeptide having a P450 activity.
An isolated, synthetic or recombinant nucleic acid comprising a nucleic acid sequence having at least 95% sequence identity to SEQ ID NO:23, wherein the nucleic acid encodes a polypeptide having epoxidation activity and has a sequence comprising a variation of SEQ ID NO:24, wherein one or more amino acid residues of SEQ ID NO:24 are replaced with a conservative substitution.
An isolated, synthetic or recombinant nucleic acid comprising a polypeptide coding sequence having at least 95% sequence identity to SEQ ID NO:23, wherein the nucleic acid encodes a polypeptide having epoxidation activity, and a bacterial, a viral or a eukaryotic promoter, wherein the coding sequence is operably linked to the promoter.
An isolated, synthetic or recombinant nucleic acid comprising a nucleic acid sequence having at least 95% sequence identity to SEQ ID NO:23, wherein the nucleic acid encodes a polypeptide having epoxidation activity, and a nucleic acid sequence encoding a detection or a purification domain.
An isolated, synthetic or recombinant nucleic acid comprising a nucleic acid sequence having at least 95% sequence identity to SEQ ID NO:23, wherein the nucleic acid encodes a polypeptide having epoxidation activity.
An isolated, synthetic or recombinant nucleic acid comprising a nucleic acid sequence having at least 98% sequence identity to SEQ ID NO:23, wherein the nucleic acid encodes a polypeptide having epoxidation activity.
An isolated, synthetic or recombinant nucleic acid comprising a nucleic acid sequence having at least 99% sequence identity to SEQ ID NO:23, wherein the nucleic acid encodes a polypeptide having epoxidation activity.
An isolated, synthetic or recombinant nucleic acid, wherein the nucleic acid comprises the a nucleic acid sequence of SEQ ID NO:23, wherein the nucleic acid encodes a polypeptide having a P450 activity and the P450 activity comprises an epoxidation activity.
Dependencies are inferred from the claim text.