On the Effects of Thiocyanate and Venturicidin on Respiration-Driven Proton Translocation in Paracoccus Denitrificans

Biochim Biophys Acta. 1984 Jul 27;766(1):222-32. doi: 10.1016/0005-2728(84)90235-4.

Abstract

A fast-responding O2 electrode has been used to confirm and extend observations of a significant kinetic discrepancy between O2 reduction and consequent proton translocation in 'O2-pulse' experiments in intact cells of P. denitrificans. The permeant, chaotropic SCN- ion abolishes this discrepancy, and greatly increases the observable----H+/O ratio, to a value approaching its accepted, true, limiting stoichiometry. The observable H+ decay rates are very slow, particularly in the absence of SCN-. The submaximal----H+/O ratios observed in the absence of SCN- are essentially independent of the size of the O2 pulse, in a manner not easily explained by a delocalised chemiosmotic energy-coupling scheme. Osmotically active protoplasts of P. denitrificans do not show a significant kinetic discrepancy between O2 reduction and H+ translocation, even in the the absence of SCN-. However, the submaximal----H+/O ratios observed in the absence of SCN- are again essentially independent of the size of the O2 pulse. As in intact cells, the observable H+ decay rates are very slow. The energy-transfer inhibitor venturicidin causes a significant increase in the----H+/O ratio observed in protoplasts of P. denitrificans in the absence of SCN-; the decay kinetics of the H+ translocation process are also somewhat modified. Nevertheless, the----H+/O ratio observed in the presence of venturicidin is also independent of the size of the O2 pulse. This observation militates further against arguments in which (a) a non-ohmic leak of protons from the bulk aqueous phase might alone be the cause of the low----H+/O ratios observed in the absence of SCN-, and (b) in which there might be a delta p-dependent change ('redox slip') in the actual----H+/O ratio. It is concluded that the observable protonmotive activity of the respiratory chain of P. denitrificans in the absence of SCN- is directly influenced by the state of the H+-ATP synthetase in the cytoplasmic membrane of this organism. We are unable to explain the data in terms of a model in which the putative protonmotive force may be acting to affect the----H+/O ratio. The possibility is considered that the delocalised bulk-to-bulk phase membrane potential set up in response to protonmotive activity is energetically insignificant.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Cell Membrane / drug effects
  • Cell Membrane / metabolism
  • Hydrogen-Ion Concentration
  • Kinetics
  • Lactones / pharmacology*
  • Oxygen Consumption / drug effects*
  • Paracoccus denitrificans / drug effects
  • Paracoccus denitrificans / metabolism*
  • Thiocyanates / pharmacology*
  • Venturicidins / pharmacology*

Substances

  • Lactones
  • Thiocyanates
  • Venturicidins