Enterobacter cloacae NAD(P)H:nitroreductase (NR; EC 18.104.22.168) catalyzes two-electron reduction of a series of quinoidal compounds according to a "ping-pong" scheme, with marked substrate inhibition by quinones. The steady-state catalytic constants (kcat) range from 0.1 to 1600 s-1, and bimolecular rate constants (kcat/Km) range from 103 to 108 M-1 s-1. Quinones, nitroaromatic compounds and competitive to NADH inhibitor dicumarol, quench the flavin mononucleotide (FMN) fluorescence of nitroreductase. The reactivity of NR with single-electron acceptors is consistent with an "outer-sphere" electron transfer model, taking into account high potential of FMN semiquinone/FMNH- couple and good solvent accessibility of FMN. However, the single-electron acceptor 1,1′-dibenzyl-4,4′-bipyridinium was far less reactive than quinones possessing similar single-electron reduction potentials (E71). For all quinoidal compounds except 2-hydroxy-1,4-naphthoquinones, there existed parabolic correlations between the log of rate constants of quinone reduction and their E71 or hydride-transfer potential (E7(Q/QH-)). Based on pH dependence of rate constants, a single-step hydride transfer seems to be a more feasible quinone reduction mechanism. The reactivities of 2-hydroxy-1,4-naphthoquinones were much higher than expected from their reduction potential. Most probably, their enhanced reactivity was determined by their binding at or close to the binding site of NADH and dicumarol, whereas other quinones used the alternative, currently unidentified binding site.
|Number of pages||10|
|Journal||Archives of Biochemistry and Biophysics|
|State||Published - Jul 15 2002|
Bibliographical noteFunding Information:
The work was supported in part by the Lithuanian State Science and Studies Foundation, and by the Petroleum Research Foundation.
- Electron transfer
ASJC Scopus subject areas
- Molecular Biology