Kinetic and Binding Properties of the Oxoglutarate Translocator of Rat-Heart Mitochondria

Abstract

The kinetic [K] study of the oxoglutarateout/malatein exchange through the inner membrane of rat-heart mitochondria was extended to higher external-oxoglutarate and to lower internal-malate concentrations. External oxoglutarate inhibits the exchange at high concentration. This excess-substrate inhibition is preceded by 4 jumps. The K-saturation curve by the internal malate presents an apparent positive cooperativity that may be interpreted in different ways. The independence of the effects of the 2 substrates on the initial rate was observed. A method for the determination of oxoglutarate binding to the external face of the inner membrane is described. The binding curve shows 4 intermediary plateau regions that reflect significant apparent K-effects, alternatively negative and positive. For external-oxoglutarate concentrations below the region of excess-substrate inhibition, the binding-saturation curve and the K-saturation curve are similar, demonstrating that K-effects are predominant. A particularly wide intermediary plateau that seems to correspond to half saturation of the active sites is common to both saturation curves. A clear lack of proportionality between the 2 curves at low oxoglutarate concentrations seems to indicate that more than 1 catalytic-rate constant is implied in the exchange kinetics. Models of the oxoglutarate carrier are presented which lead to a minimum degree of 10:10 for the equation of the binding of oxoglutarate to the catalytic sites. In the 1st model this corresponds to 10 subunits associated into a single oligomer while in the 2nd model this results from a mixture of monomeric, dimeric, trimeric and tetrameric associations.