Compartmented coupling of chicken heart mitochondrial creatine kinase to the nucleotide translocase requires the outer mitochondrial membrane.
The kinetic coupling of mitochondrial creatine kinase (MiMi-CK) to ADP/ATP translocase in chicken heart mitochondrial preparations is demonstrated. Measuring the MiMi-CK apparent Km value for MgATP2- (at saturating creatine) gives a value of 36 microM when MiMi-CK is coupled to oxidative phosphorylation. This Km value is threefold lower than the Km for enzyme bound to mitoplasts or free in solution. The nucleotide translocase Km value for ADP decreases from 20 to 10 microM in the presence of 50 mM creatine only with intact mitochondria. Similar experiments with mitoplasts do not give decreased Km values. The observed Km differences can be used to calculate the concentration of ATP and ADP under steady-state conditions showing that the observed differences in the kinetic constants accurately reflect the enzyme activities of MiMi-CK under the different conditions. The behavior of the Km values provides evidence for what we term compartmented coupling. Therefore, like the rabbit heart system (S. Erickson-Viitanen, P. Viitanen, P. J. Geiger, W. C. T. Yang, and S. P. Bessman (1982) J. Biol. Chem. 257, 14395-14404) compartmented coupling requires an intact outer mitochondrial membrane. The apparent Km values for normal or compartmentally coupled systems can be used to calculate steady-state values of ATP and ADP by coupling enzyme theory. Hence, the overall kinetic parameters accurately reflect the behavior of the enzymes whether free in solution or in the intermembrane space.
Brooks
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Suelter
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CH
D000244:Adenosine Diphosphate / Q000032:analysis; D000818:Animals; D002451:Cell Compartmentation; D002462:Cell Membrane / Q000502:physiology; D002645:Chickens; D003402:Creatine Kinase / Q000032:analysis*; D007700:Kinetics; D008929:Mitochondria, Heart / Q000201:enzymology*; D000226:Mitochondrial ADP, ATP Translocases / Q000032:analysis*; D009713:Nucleotidyltransferases / Q000032:analysis*; D010085:Oxidative Phosphorylation; D010725:Phosphocreatine / Q000096:biosynthesis
D010725:Phosphocreatine; D000244:Adenosine Diphosphate; D000226:Mitochondrial ADP, ATP Translocases; D003402:Creatine Kinase; D009713:Nucleotidyltransferases