[Affinity modification of creatine kinase and ATP-ADP translocase in heart mitochondria: determination of their stoichiometric ratio].
The interaction of mitochondrial creatine kinase and ATP-ADP translocase with 2.3-dialdehyde derivatives of ADP and ATP (oADP and oATP) has been studied. It was shown that these compounds are irreversible and specific inhibitors of creatine kinase (KioADP = 0.6mM, KioATP = 1.12 mM) and ATP-ADP translocase (KioADP = 0.065mM, KioATP = 0.14 mM). The substrates protect both enzymes from inactivation by these compounds. The maximal pseudo-first order rate constants for the 2,3-dialdehyde nucleotide derivative interaction with creatine kinase are 0.2 min-1 for oADP (pH 6.5) and 0.11 min-1 for oATP (pH 7.0). A decrease in the creatine kinase activity correlates with the incorporation of the reagent into the protein. The completely inactivated, isolated and purified enzyme contains 1 mol of oADP per mole of active sites. A procedure for simultaneous determination of the creatine kinase and translocase content in mitochondria and mitoplasts has been developed, which is based on the application of [3H]oADP in combination with specific treatment of mitochondria (or mitoplasts) with carboxyatractyloside 2,4-dinitrofluorobenzene and a mixture of creatine kinase substrates (MgADP + phosphocreatine). It has been found that for heart mitochondria from different animals the content of creatine kinase and translocase is 2.1-2.6 and 2.4-2.9 mol per mol of cytochrome c oxidase, respectively. Thus, the stoiochiometric ratio of creatine kinase and ATP-ADP translocase is close to 1.0 for all mitochondrial preparations under study (i.e. rat, dog, rabbit and chicken).
D000244:Adenosine Diphosphate / Q000031:analogs & derivatives / Q000378:metabolism / Q000494:pharmacology; D000255:Adenosine Triphosphate / Q000031:analogs & derivatives / Q000494:pharmacology; D000818:Animals; D003402:Creatine Kinase / Q000032:analysis / Q000037:antagonists & inhibitors*; D007425:Intracellular Membranes / Q000201:enzymology; D007700:Kinetics; D008929:Mitochondria, Heart / Q000201:enzymology*; D000226:Mitochondrial ADP, ATP Translocases / Q000032:analysis / Q000037:antagonists & inhibitors*; D009713:Nucleotidyltransferases / Q000037:antagonists & inhibitors*; D051381:Rats
C017199:2',3'-dialdehyde ATP; D000244:Adenosine Diphosphate; C018808:adenosine 5'-diphosphate 2',3'-dialdehyde; D000255:Adenosine Triphosphate; D000226:Mitochondrial ADP, ATP Translocases; D003402:Creatine Kinase; D009713:Nucleotidyltransferases
Affinnaia modifikatsiia kreatinkinazy i ATP-ADP translokazy v mitokhondriiakh serdtsa, opredelenie ikh stekhiometricheskogo sootnosheniia.