Molecular characterization of two genes from Streptomyces tendae Tu901 required for the formation of the 4-formyl-4-imidazolin-2-one-containing nucleoside moiety of the peptidyl nucleoside antibiotic nikkomycin

The genes nikQ and nikR were identified by sequencing DNA of the nikkomycin biosynthetic gene cluster from Streptomyces tendae Tu901/8c. The nikQ gene encodes a P450 cytochrome, and the predicted NikR gene product shows 48-56 % sequence identity with uracil phosphoribosyltransferases from eukaryotic organisms. The nikQ and nikR genes were inactivated separately by insertion of a kanamycin-resistance cassette. Inactivation of the nikQ gene abolishe d synthesis of nikkomycins containing 4-formyl-4-imidazolin-2-one as the ba se (nikkomycins X and I), whereas production of nikkomycins containing urac il (nikkomycins Z and J) was not affected. Nikkomycin X and I production co uld be restored by feeding 4-formyl-4-imidazolin-2-one to the nikQ mutants, indicating that NikQ is responsible for its formation from l-histidine. Di sruption of the nikR gene resulted in formation of decreased amounts of nik komycins X and I, whereas nikkomycins Z and J were synthesized at wild-type levels. A fluorouracil-resistant nikR mutant lacking uracil phosphoribosyl transferase (UPRTase) activity did not synthesize nikkomycins X and I and a ccumulated 4-formyl-4-imidazolin-2-one in its culture filtrate, whereas for mation of nikkomycins Z and J was unimpaired. The mutant was complemented t o nikkomycin X and I production by nikR expressed from the mel promoter of plasmid pIJ702. The nikR gene expressed in Escherichia coli led to the prod uction of UPRTase activity. Our results indicate that NikR converts 4-formy l-4-imidazolin-2-one to yield 5'-phosphoribosyl-4-formyl-4-imidazolin-2-one , the precursor of nikkomycins containing this base.