| The inner mitochondrial membrane contains transport systems catalyzing influx and efflux of cations between the cytosol and mitochondrial matrix. Our laboratory studies three distinct cation cycles, and each of these cycles plays a major physiological role in the overall energy economy.; The Ca2+ cycle is responsible for rapid oscillations in matrix Ca2+, which result in sustained activation of Ca 2+-sensitive dehydrogenases and regulation of ATP production. The efflux of Ca2+ from the mitochondrial matrix is carried by the Na+/Ca2+ antiporter. We purified the Na +/Ca2+ antiporter from beef heart mitochondria and reconstituted the antiporter into liposomes. The kinetics of the Na +/Ca2+ exchange were consistent with its participation in rapid Ca2+ oscillation. The reconstitution also revealed that the antiporter was capable of both electroneutral and electrophoretic Na+/Ca2+ exchange, the mode of transport depending on the availability of a pathway for charge-compensating ion transport.; The K+ cycle maintains the integrity of the vesicular structure of the inner membrane. Influx of K+ into the matrix is catalyzed by the mitochondrial KATP channel (mitoK ATP). MitoKATP is hypothesized to be the receptor for the cardioprotective effects of K+ channel openers (KCO) and for blocking the cardioprotection by glyburide and 5-hydroxydecanoate (5-HD). We have studied the effect of glyburide and 5-HD on isolated, respiring mitochondria. Our results show that glyburide and 5-HD are potent blockers of K+ flux through mitoKATP only in open states, in which Mg 2+, ATP, and physiological (GTP) or pharmacological (KCO) openers are present. These results are consistent with a role for mitoKATP in cardioprotection.; The H+ cycle has been previously characterized only in brown adipose tissue mitochondria and consists of uncoupling protein (UCP1), which dissipates energy and generates heat by catalyzing back-flux of protons into the mitochondrial matrix. Recently, proteins homologous to UCP1 were discovered in many other tissues, including white fat and skeletal muscle. If the newly discovered UCP2 and UCP3 function similarly, they will enhance peripheral energy expenditure and are potential targets for the treatment of obesity. We have reconstituted bacterially expressed UCP2 and UCP3 into liposomes and shown that UCP2 and UCP3 behave similarly to UCP1, i.e., they catalyze electrophoretic flux of protons and alkylsulfonates, and proton flux exhibits an obligatory requirement for fatty acids. Proton flux is inhibited by purine nucleotides. These findings are consistent with the hypothesis that UCP2 and UCP3 behave as uncoupling proteins in the cell. |
