Nikkomycin biosynthesis: formation of a 4-electron oxidation product during turnover of NikD with its physiological substrate

Nikkomycins are peptidyl nucleoside antibiotics that act as therapeutic antifungal agents in humans and easily degraded insecticides in agriculture. The nikkomycin peptidyl moiety contains a pyridyl residue derived from L-lysine. The first step in peptidyl biosynthesis is an aminotransferase-catalyzed reaction that converts L-lysine to Delta(1)- or Delta(2)-piperideine-2-carboxylate (P2C). Spectral, chromatographic, and kinetic analyses show that the aerobic reaction of nikD with P2C results in the stoichiometric formation of picolinate, accompanied by the reduction of 2 mol of oxygen to hydrogen peroxide. A high resolution HPLC method, capable of separating picolinate, nicotinate and isonicotinate, was developed and used in product identification. NikD contains 1 mol of covalently bound FAD and exists as a monomer in solution. Reductive and oxidative titrations with dithionite and potassium ferricyanide, respectively, show that FAD is the only redox-active group in nikD. Anaerobic reaction of nikD with 1 mol of P2C results in immediate reduction of enzyme-bound FAD. Because nikD is an obligate 2-electron acceptor, it is proposed that the observed 4-electron oxidation of P2C to picolinate occurs via a mechanism involving two successive nikD-catalyzed 2-electron oxidation steps. In addition to nikkomycins, a nikD-like reaction is implicated in the biosynthesis of an L-lysine-derived pyridyl moiety found in streptogramin group B antibiotics that are used as part of a last resort treatment for severe infections due to gram positive bacteria.