A novel inhibitor of NADH dehydrogenase in Paracoccus denitrificans.

In [ l ] low concentrations of Tinopal AN, (1,l-bis(3,N-5-dimethylbenzoxazol-2-y1)methine p-toluene sulphonate), which is present in the optical brightener Uvitex AN (Ciba-Geigy Ltd) [2], inhibited the aerobic respiration of Paracoccus denitrificans. Using inverted membrane vesicles of P. denitrifcans [I], this inhibition was shown to occur in the NADH dehydrogenase (EC 1.66.99.3) region of the respiratory chain because, although respiration through segments I1 and I11 of the respiratory chain was unaffected by relatively high concentrations of the compound, it was markedly inhibited when NADH was used as the electron donor. The sensitivity of growing cells of P, denitrificans to rotenone, a well-known segment I inhibitor, has been ascribed [3] to the possession of iron-sulphur centre N-2 in the NADH dehydrogenase complex of the respiratory chain. This iron-sulphur centre was absent when rotenone-insensitive cells ofP. denitrificans were examined by electron paramagnetic resonance spectroscopy [3]. When amytal and piericidin A (two other classical inhibitors of the NADH dehydrogenase region of the respiratory chain) were compared to rotenone in beef-heart submitochondrial particles, it was concluded [4] that both amytal and piericidin A bound to the same site as rotenone, since they competed with [14C]rotenone for binding at the rotenonebinding site. It was thus deduced that all 3 inhibitors share the same binding site. We report here, by comparison between rotenonesensitive and rotenone-insensitive cells of P. denitrificnns, that Tinopal AN did not share the same specific site as rotenone (and also, presumably, arnytal and piericidin A), and therefore provides the first example of an inhibitor of the NADH dehydrogenase region of the respiratory chain whose binding site is not associated with the iron-sulphur centre N-2.