P450-BM3 variants with improved activity
The present invention provides improved P450-BM3 variants with improved activity. In some embodiments, the P450-BM3 variants exhibit improved activity over a wide range of substrates.
1. A recombinant cytochrome P450-BM3 variant having at least 90% sequence identity to a polypeptide sequence comprising the sequence set forth in SEQ ID NO:2.
2. The recombinant cytochrome P450-BM3 variant of claim 1 , wherein said variant oxidizes at least three organic substrates.
3. The recombinant cytochrome P450-BM3 variant of claim 2 , wherein said organic substrate is selected from loratadine, imatinib, geftinib, and diclofenac.
4. A recombinant polynucleotide sequence encoding a recombinant cytochrome P450-BM3 variant having at least 90% sequence identity to a polynucleotide sequence comprising the sequence set forth in SEQ ID NO: 1.
5. A recombinant polynucleotide sequence encoding a recombinant cytochrome P450-BM3 variant of claim 1 .
6. The recombinant polynucleotide sequence of claim 4 , wherein said sequence comprises SEQ ID NO: 1.
7. An expression vector comprising the polynucleotide sequence of claim 4 .
8. The vector of claim 7 , wherein said polynucleotide sequence is operably linked with regulatory sequences suitable for expression of said polynucleotide sequence in a suitable host cell.
9. The vector of claim 8 , wherein said host cell is a prokaryotic or eukaryotic cell.
10. The vector of claim 9 , wherein said host cell is a prokaryotic cell.
11. The vector of claim 10 , wherein said host cell is E. coli.
12. A host cell comprising the vector of claim 7 .
13. A method for producing at least one recombinant cytochrome P450-BM3 variant comprising culturing a host cell under conditions such that at least one recombinant cytochrome P450-BM3 variant of claim 1 is produced by said host cell.
14. The method of claim 13 , further comprising the step of recovering said at least one recombinant cytochrome P450-BM3 variant.