Molecular characterization of co-transcribed genes from Streptomyces tendae Tü901 involved in the biosynthesis of the peptidyl moiety and assembly of the peptidyl nucleoside antibiotic nikkomycin

Abstract

Six genes (nikP1, nikP2, nikS, nikT, nikU, and nikV) from Streptomyces tendae Tu901 were identified by analysis of the nucleotide sequence of the nikkomycin gene cluster. These genes, together with the previously described nikQ 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 peptide synthase consisting of an adenylation domain, a thiolation domain, and an N-terminal 70-residue segment of unknown function. The amino acid sequence encoded by the nikP2 gene displays similarity to the sequences of thioesterases, and the nikS product belongs to a superfamily of proteins characterized by a specific ATP-binding fold. The N-terminal 70 amino acids of the predicted nikT gene product show significant sequence similarity to acyl carrier proteins, and the C-terminal 330 amino acids to aminotransferases. The sequences of the deduced proteins NikU and NikV exhibit similarity to components S and E, respectively, of glutamate mutase from Clostridium. Disruption of the nikP1, nikS, nikT, or nikV gene by insertion of a kanamycin resistance cassette abolished formation of nikkomycins I, J, X, and Z, all of which contain hydroxypyridylhomothreonine as the peptidyl moiety. The nikP1 mutants, and the nikS and nikT mutants accumulated the nucleoside moieties nikkomycin Cz, and nikkomycins Cx and Cz, respectively. The nikV mutants formed nikkomycins Ox and Oz, which contain 2-amino-4-hydroxy-4-(3'-hydroxy-6'-pyridyl) butanoic acid as the peptidyl moiety. The nikP2 mutants synthesized nikkomycins I, J, X, and Z, but amounts of nikkomycins I and X, which contain formylimidazolone as the base, were lower. Feeding formylimidazolone 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 hydroxypyridylhomothreonine, the genes nikP1, nikP2 and nikS are required for the assembly of nikkomycins, and nikT is required for both pathways. The putative activities of each of their products are discussed.

Structural and Functional Characterization of NikO, an Enolpyruvyl Transferase Essential in Nikkomycin Biosynthesis

“…Their biosynthesis is complex and proceeds via two separate pathways producing hydroxypyridyl-homothreonine and amino hexuronic acid, which are joined by a peptide bond in a final step (43,44). The enzyme catalyzing the first step in the formation of the aminohexuronic acid moiety, NikO, belongs to the Structural Classification of Proteins (SCOP) superfamily of enolpyruvyl transferases (SCOP number: 55209).…”

The 1.92-Å Structure of Streptomyces coelicolor A3(2) CYP154C1

“…The hydroxyl or epoxide substituents provide an important layer of structural variability into the final natural product structures and often significantly influence biological activity (12,22). P450 monooxygenases are also involved in one of the initial steps in formation of the coumarin group of antibiotics (31), and of the peptidyl nucleoside antibiotic nikkomycin (32), as well as in oxidative tailoring of the vancomycin-like glycopeptides balhimycin (16) and complestatin (33). Ultimately, the power to manipulate macrolide metabolic systems using combinatorial biosynthetic technology (34 -36) will be extended by identification and/or engineering of additional monooxygenases with versatile activities to provide novel biologically active natural products.…”

Amino Acid Precursor Supply in the Biosynthesis of the RNA Polymerase Inhibitor Streptolydigin by Streptomyces lydicus

“…SlgE1 and SlgE2 show significant similarities to VinH and VinI, respectively, S and E subunits of coenzyme B12 (adenosylcobalamin)-dependent mutase from S. halstedii (19). They are also similar to the pairs NikU-NikV, SanU-SanV, and GlmA-GlmB, S and E subunits of GMs from S. tendae, S. ansochromogenes, and Actinoplanes friuliensis involved in the biosynthesis of nikkomycin (13,14) and friulimycin (10), respectively. The participation of slgE1 and slgE2 in streptolydigin biosynthesis was assessed by the simultaneous deletion of both genes from S. lydicus, thus generating mutant SLME1E2 (Fig.…”