Na+ is translocated at NADH:quinone oxidoreductase segment in the respiratory chain of Vibrio alginolyticus.

H Tokuda,T Unemoto

doi:10.1016/s0021-9258(17)42862-6

Abstract

The coupling site of the Na+ pump to the respiratory chain of Vibrio alginolyticus was examined using membrane fractions prepared from the wild type, Na+ pump-deficient mutants, and spontaneous revertant. NADH oxidase of the wild type and revertant specifically required NA+ for maximum activity, whereas Na+ was not essential for the NADH oxidase of mutants. Similar to the Na+ pump in whole cells, the Na+-dependent NADH oxidase in membranes had a pH optimum in the alkaline region. A respiratory inhibitor, 2-heptyl-4-hydroxyquinoline-N-oxide (HQNO), inhibited the Na+-dependent NADH oxidase but had little effect on the NA+-independent activity of mutant membranes. NADH:quinone oxidoreductase was found to be the Na+-dependent HQNO-sensitive site of the NADH oxidase. In the wild type cells, HQNO was also found to cause a strong inhibition of the Na+ pump with little effect on the overall H+ extrusion by respiration. The inhibition of the Na+ pump by HQNO was overcome by oxidized, but not reduced, N,N,N',N'-tetra-methyl-p-phenylenediamine (TMPD). In the presence of oxidised TMPD, the electron flow NADH to oxygen seemed to bypass the HQNO-sensitive site and energize the Na+ pump. From these results, it was concluded that the Na+ pump is coupled to the respiratory chain at the step of NADH:quinone oxidoreductase.

Highlights

The coupling site of the Nap+ump to therespiratory uptake of solutes (1)and growth itself (3) is insensitive chain of Vibrioalginolyticus wasexaminedusing to CCCP when the Na' pump functions
Na' Requirement of NADH Oxidase-Effect of salts on NADH oxidase activity was examined with membrane fractions isolated from various strains of V. alginolyticus grown on the synthetic medium containing glycerol
It is notable that the G3P-linked QH2 formation by the wild type membranes was neither dependent on Na+ (Table I) nor sensitive to HQNO. These results indicate that HQNO inhibits the Na+dependent site of NADH oxidase, that is, the Na'-dependent QH2formation by the NADH:quinone oxidoreductase

Summary

Introduction

The coupling site of the Nap+ump to therespiratory uptake of solutes (1)and growth itself (3) is insensitive chain of Vibrioalginolyticus wasexaminedusing to CCCP when the Na' pump functions. As promembrane fractions prepared from the wild type,Na* posed by the chemiosmotic theory of Mitchell (4, 5), the pump-deficient mutants, and spontaneous revertant. Respiratory inhibitor, 2heptyl-4-hydroxyquinoline-N-oxide(HQNO), inhibited the Na+-dependent NADH oxidase but had little effect on the Na+-independent activity ofmutant membranes. In the wild type cells,HQNO was electron-carrying component alternate in thredox loop, profound to causea strong inhibition of the Na+pump with tonations and deprotonationosccur on opposite sidesof memlittle effecton the overallH+extrusion by respiration. In the presence that the extrusion of H+ is a separate process from electron of oxidised TMPD, the electron flow from NADH to transport andperformed through the H+-conducting channel oxygen seemed to bypass the HQNO-sensitive site and of redox proteins which is energized by electrontransfer

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Conclusion