Nature of proton cycling during gramicidin uncoupling of oxidative phosphorylation.
Addition of gramicidin D to liver mitochondria, incubated in low- or high-salt media, results in stimulation of respiration in the absence or presence of depression of delta muH, respectively. Gramicidin D concentrations 2 orders of magnitude higher are required in the low-salt media with full uncoupling at 1 nmol of gramicidin.mg-1. The stimulation of respiration is not accompanied by increased passive proton influx in low-salt media. In high-salt media, the extent of respiratory stimulation and the extent of delta muH depression differ according to the nature and concentration of cation. The flow-force relationship is very steep when gramicidin D induced uncoupling occurs in low-salt media and much less steep in high-salt media. A multiplicity of flow-force relationship, respiratory rate vs delta muH, is obtained, the slope of which depends on the nature and concentration of cation, and which can be reproduced by computer simulation by introducing a variable extent of proton cycling either in the membrane or in the pump. The apparent proton conductance, as analyzed in the relationship of Je/delta muH vs delta muH, increases in the so-called ohmic and nonohmic regions according to whether gramicidin D is added in high-salt or low-salt media, respectively. Titration with antimycin of the respiratory control ratio (RCR) in gramicidin D treated mitochondria leads to a depression of the RCR in high-salt but not in low-salt media. The view is discussed that in low-salt media the gramicidin D induced uncoupling is due to a cycling of protons within a proton domain operationally located at or near the proton pump.(ABSTRACT TRUNCATED AT 250 WORDS)
Luvisetto
S
S
CNR Unit for the Study of the Physiology of Mitochondria, University of Padova, Italy.
Azzone
G F
GF
D000818:Animals; D002259:Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone / Q000494:pharmacology; D006096:Gramicidin / Q000494:pharmacology*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008433:Mathematics; D008930:Mitochondria, Liver / Q000187:drug effects / Q000378:metabolism*; D008962:Models, Theoretical; D010085:Oxidative Phosphorylation / Q000187:drug effects*; D010101:Oxygen Consumption / Q000187:drug effects*; D011522:Protons; D051381:Rats; D014475:Uncoupling Agents
D011522:Protons; D014475:Uncoupling Agents; D006096:Gramicidin; D002259:Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone