Deletion of rapQONML from the rapamycin gene cluster of Streptomyces hygroscopicus gives production of the 16-O-desmethyl-27-desmethoxy analog.
Five contiguous genes in the rapamycin gene cluster, rapQONML, of Streptomyces hygroscopicus ATCC29253 were replaced with a neomycin resistance marker by double homologous recombination. The resulting strain, if fed pipecolate, produced the analog 16-O-desmethyl-27-desmethoxyrapamycin instead of rapamycin. This indicates that the P450 hydroxylase encoded by rapN is specific for C-27, and that the O-methyltransferases encoded by rapQ and rapM methylate the hydroxyl groups on C-16 and C-27. By inference, the remaining P450 hydroxylase and methyltransferase genes (rapI and rapJ) are responsible for hydroxylation of C-9 and methylation of the C-39 hydroxyl, consistent with their homology to fkbD and fkbM, respectively, in the FK506 cluster. The relatively high level of 16-O-desmethyl-27-desmethoxyrapamycin produced indicates that the reactions at C-9 and C-39 do not require previous modification of the macrolactone precursor at either C-16 or C-27.
Chung
L
L
Kosan Biosciences, Inc, Hayward, CA 94545, USA.
Liu
L
L
Patel
S
S
Carney
J R
JR
Reeves
C D
CD
D000900:Anti-Bacterial Agents / Q000096:biosynthesis* / Q000737:chemistry / Q000378:metabolism*; D001483:Base Sequence; D017931:DNA Primers / Q000235:genetics; D017353:Gene Deletion; D005798:Genes, Bacterial; D005818:Genetic Engineering; D006900:Hydroxylation; D018942:Macrolides; D008745:Methylation; D015394:Molecular Structure; D005810:Multigene Family; D020123:Sirolimus / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism; D013302:Streptomyces / Q000235:genetics* / Q000378:metabolism*
C427742:16-O-desmethyl-17-desmethoxyrapamycin; D000900:Anti-Bacterial Agents; D017931:DNA Primers; D018942:Macrolides; D020123:Sirolimus