linoleic-acid and benzohydroxamic-acid

In this study we demonstrated that mitochondria of Candida parapsilosis contain a constitutive ubiquinol alternative oxidase (AOX) in addition to a classical respiratory chain (CRC) and a parallel respiratory chain (PAR) both terminating by two different cytochrome c oxidases. The C. parapsilosis AOX is characterized by a fungi-type regulation by GMP (as a stimulator) and linoleic acid (as an inhibitor). Inhibitor screening of the respiratory network by the ADP/O ratio and state 3 respiration determinations showed that (i) oxygen can be reduced by the three terminal oxidases through four paths implying one bypass between CRC and PAR and (ii) the sum of CRC, AOX and PAR capacities is higher than the overall respiration (no additivity) and that their engagement could be progressive according to the redox state of ubiquinone, i.e. first cytochrome pathway, then AOX and finally PAR.

Topics: Adenosine Diphosphate; Candida; Cell Respiration; Cyanates; Electron Transport; Guanosine Monophosphate; Hydroxamic Acids; Linoleic Acid; Mitochondria; Mitochondrial Proteins; NAD; Oxidoreductases; Oxygen; Plant Proteins

An uncoupling protein was recently discovered in plant mitochondria and demonstrated to function similarly to the uncoupling protein of brown adipose tissue. In this work, green tomato fruit mitochondria were purified on a self-generating Percoll gradient in the presence of 0.5% bovine serum albumin to deplete mitochondria of endogenous free fatty acids. The uncoupling protein activity was induced by the addition of linoleic acid during the resting state, and in the progressively uncoupled state, as well as during phosphorylating respiration in the presence of benzohydroxamic acid, an inhibitor of the alternative oxidase and with succinate (+ rotenone) as oxidizable substrate. Linoleic acid strongly stimulated the resting respiration in fatty acid-depleted mitochondria but had no effect on phosphorylating respiration, suggesting no activity of the uncoupling protein in this respiratory state. Progressive uncoupling of state 4 respiration decreased the stimulation by linoleic acid. The similar respiratory rates in phosphorylating and fully uncoupled respiration in the presence and absence of linoleic acid suggested that a rate-limiting step on the dehydrogenase side of the respiratory chain was responsible for the insensitivity of phosphorylating respiration to linoleic acid. Indeed, the ADP/O ratio determined by ADP/O pulse method was decreased by linoleic acid, indicating that uncoupling protein was active during phosphorylating respiration and was able to divert energy from oxidative phosphorylation. Moreover, the respiration rates appeared to be determined by membrane potential independently of the presence of linoleic acid, indicating that linoleic acid-induced stimulation of respiration is due to a pure protonophoric activity without any direct effect on the electron transport chain.

Topics: Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone; Carrier Proteins; Hydroxamic Acids; Ion Channels; Linoleic Acid; Membrane Potentials; Membrane Proteins; Mitochondria; Mitochondrial Proteins; Oxidative Phosphorylation; Oxygen Consumption; Phosphorylation; Plant Proteins; Solanum lycopersicum; Uncoupling Agents; Uncoupling Protein 1