Genes and proteins for the biosynthesis of rosaramicin
OwnerChris M. Farnet, Alfredo Staffa +1inventors
InventorsChris M. Farnet, Alfredo Staffa, Xianshu Yang
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Abstract
Genes and proteins involved in the biosynthesis of macrolides by microorganisms, in particular the nucleic acids forming the biosynthetic locus for the 16-member macrolide rosaramicin from Micromonospora carbonacea. These nucleic acids can be used to make expression constructs and transformed host cells for the production of rosaramicin. The genes and proteins allow direct manipulation of macrolides and related chemical structures via chemical engineering of the proteins involved in the biosynthesis of rosaramicin.
Claims28
An isolated, purified or enriched nucleic acid comprising a nucleic acid sequence selected from the group consisting of:
- (a) a nucleic acid of SEQ ID NOS:
- 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31, 33, 35, 37, 39;
- (b) a nucleic acid encoding a polypeptide of SEQ ID NOS:
- 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, 30, 32, 34, 36, 38;
- (c) a nucleic acid having at least 75% homology to a nucleic acid of (a) or (b) as determined by analysis with BLASTN version 2.0 with the default parameters;
- (d) a nucleic acid complementary to a nucleic acid of (a), (b) or (c).
An isolated, purified or enriched nucleic acid comprising a nucleic acid that hybridizes under stringent conditions to any one of rosaramicin open reading frames (ORFs) 1 to 19 (SEQ ID NOS:
- 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31, 33, 35, 37, 39) and can substitute for the ORF to which it specifically hybridizes to direct the synthesis of a rosaramicin compound or analogue.
An isolated, purified or enriched nucleic acid that hybridizes under stringent conditions to any one of rosaramicin ORFs 1, 2, 4, 5, 6, 7, 8, 9, 10, 12, 14 or 15 (SEQ ID NOS:
- 3, 5, 9, 11, 13, 15, 17, 19, 21, 25, 29, and 31) and can substitute for the ORF to which it specifically hybridizes to direct the synthesis of a rosaramicin compound or analogue.
An isolated gene cluster comprising ORFs encoding polypeptides sufficient to direct the synthesis of a rosaramicin compound or analogue.
An isolated polypeptide comprising a polypeptide sequence selected from any one of:
- (a) a polypeptide of any one of SEQ ID NOS:
- 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, 30, 32, 34, 36, 38;
- and
- (b) a polypeptide which is at least 75% identical in amino acid sequence to a polypeptide of any one of SEQ ID NOS:
- 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, 30, 32, 34, 36, 38 as determined by analysis with BLASTP with the default parameters.
An isolated or purified antibody capable of specifically binding to a polypeptide having a sequence selected from the group consisting of SEQ ID NOS:
- 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, 30, 32, 34, 36, 38.
A method of making a polypeptide having a sequence selected from the group consisting of SEQ ID NOS:
- 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, 30, 32, 34, 36, 38 comprising introducing a nucleic acid encoding said polypeptide, said nucleic acid being operably linked to a promoter, into a host cell.
A method of making a rosaramicin compound or analog comprising culturing a Micromonospora carbonacea bacterium under conditions allowing for expression of rosaramicin ORFs 1 to 19 (SEQ ID NOS:
- 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31, 33, 35, 37, 39) present in the E. coli strains DH10B having accession nos. IDAC 100702-1, 100702-2 and 100702-3.
A computer readable medium having stored thereon a sequence selected from the group consisting of a nucleic acid code of SEQ ID NOS:
- 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31, 33, 35, 37, 39, 41, 43, 45, 47, 49, 51 and a polypeptide code of SEQ ID NOS:
- 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 44, 46, 48, 50.
A computer system comprising a processor and a data storage device wherein said data storage device has stored thereon a sequence selected from the group consisting of a nucleic acid code of SEQ ID NOS:
- 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31, 33, 35, 37, 39, 41, 43, 45, 47, 49, 51 and a polypeptide code of SEQ ID NOS:
- 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 44, 46, 48, 50.
Dependencies are inferred from the claim text.