MINIMAL STREPTOMYCES SP STRAIN C5 DAUNORUBICIN POLYKETIDE BIOSYNTHESIS GENES REQUIRED FOR AKLANONIC ACID BIOSYNTHESIS

The structure of the Streptomyces sp, strain C5 daunorubicin type II p olyketide synthase (PKS) gene region is different from that of other k nown type II PKS gene clusters, Directly downstream of the genes encod ing ketoacylsynthase alpha and beta (KSalpha, KSbeta) are two genes (d psC, dpsD) encoding proteins of unproven function, both absent from ot her type II PKS gene clusters, Also in contrast to other type II PKS c lusters, the gene encoding the acyl carrier protein (ACP), dpsG, is lo cated about 6.8 kbp upstream of the genes encoding the daunorubicin KS alpha and KSbeta. In this work, we demonstrate that the minimal genes required to produce aklanonic acid in heterologous hosts are dpsG (ACP ), dauI (regulatory activator), dpsA (KSalpha), dpsB (KSbeta), dpsF (a romatase), dpsE (polyketide reductase), and dauG (putative deoxyaklano nic acid oxygenase). The two unusual open reading frames, dpsC (KASIII homolog lacking a known active site) and dpsD (acyltransferase homolo g), are not required to synthesize aklanonic acid, Additionally, repla cement of dpsD or dpsCD in Streptomyces sp. strain C5 with a neomycin resistance gene (aphI) results in mutant strains that still produced a nthracyclines.