UNCOUPLING EFFECT OF NITRITE DURING DENITRIFICATION BY PSEUDOMONAS-FLUORESCENS - AN IN-VIVO P-31-NMR STUDY

In vivo P-31-NMR was used to investigate the basis for the inhibition of denitrification by nitrite accumulated endogenously by Pseudomonas fluorescens ATCC 17822 (bio-type II) at pH 7.0. Cells were immobilized in kappa-carrageenan to obtain high cell concentrations in the NMR tu be. Acetate and nitrate in two concentration ratios were supplied as e lectron donor and acceptor, respectively, to achieve different levels of nitrite accumulation. During denitrification, cells were able to ma intain a pH gradient of approximately 0.4 to 0.5 units, but when nitri te accumulation reached values approximating 27 mM the transmembrane D elta pH collapsed sharply. Nitrite stimulated the reduction rate of ni trate; furthermore, at nitrite concentrations below 1 mM, activation o f oxygen respiratory rates was observed in cells grown under aerobic c onditions. The results provide evidence for nitrite acting as a proton ophore (an uncoupler that increases the proton permeability of membran es by a shuttling mechanism). (C) 1996 John Wiley & Sons, Inc.