Molecular characterization of co-transcribed genes from Streptomyces tendae Tu901 involved in the biosynthesis of the peptidyl moiety and assembly ofthe peptidyl nucleoside antibiotic nikkomycin

Six genes (nik-P1, nikP2, nikS, nikT, nikU, and nikV) from Streptomyces ten dae Tu901 were identified by analysis of the nucleotide sequence of the nik komycin gene cluster. These genes, together with the previously described n ikQ and nikR, span 9.39 kb and are transcribed as a polycistronic mRNA in a growth-phase-dependent manner. The nikP1 gene encodes a non-ribosomal pept ide synthase consisting of an adenylation domain, a thiolation domain, and an N-terminal 70-residue segment of unknown function. The amino acid sequen ce encoded by the nikP2 gene displays similarity to the sequences of thioes terases, and the nikS product belongs to a superfamily of proteins characte rized by a specific ATP-binding fold. The N-terminal 70 amino acids of the predicted nikT gene product show significant sequence similarity to acyl ca rrier proteins, and the C-terminal 330 amino acids to aminotransferases. Th e sequences of the deduced proteins NikU and NikV exhibit similarity to com ponents S and E, respectively, of glutamate mutase from Clostridium. Disrup tion of the nikP1, nikS, nikT, or nikV gene by insertion of a kanamycin res istance cassette abolished formation of nikkomycins I, J, X, and Z, all of which contain hydroxypyridylhomothreonine as the peptidyl moiety. The nikP1 mutant?;, and the nikS and nikT mutants accumulated the nucleoside moietie s nikkomycin C-z, and nikkomycins C-x and C-y, respectively. The nikV mutan ts formed nikkomycins O-x and O-z, which contain 2-amino-4-hydroxy-4-(3'-hy droxy-6'-pyridyl) butanoic acid as the peptidyl moiety. The nikP2 mutants s ynthesized nikkomycins I, J, X, and Z, but amounts of nikkomycins I and X, which contain formylimidazolone as the base, were lower. Feeding formylimid azolone to nikP2 mutants restored the ability to form nikkomycins I and X. Our results indicate that nikU and nikV are required for the synthesis of h ydroxypyridylhomothreonine, the genes nikP1, nikP2 and nikS are required fo r the assembly of nikkomycins, and,nikT is required for both pathways. The putative activities of each of their products are discussed.