Interaction of creatine kinase with phosphorylating rabbit heart mitochondria and mitoplasts |
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| Authors: | C Vial O Marcillat D Goldschmidt B Font D Eichenberger |
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| Affiliation: | 1. The Key Laboratory of Reproductive Genetics, Ministry of Education (Zhejiang University), Hangzhou, China;2. The International Peace Maternity and Child Health Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, China;3. Department of Pathology and Pathophysiology, School of Medicine, Zhejiang University, Hangzhou, China;4. Bio-X Institutes, Key Laboratory for the Genetics of Developmental and Neuropsychiatric Disorders (Ministry of Education), Shanghai Jiao Tong University, Shanghai, China;1. Guangdong Provincial Key Laboratory of Marine Biology, Shantou University, Shantou 515063, China;2. College of Marine Science, South China Agriculture University, Guangzhou 510642, China;1. Department of Anesthesiology, Duke Perioperative Genomics Program, Duke University Medical Center, Durham, NC 27710, USA;2. Department of Biostatistics and Bioinformatics, Duke University Medical Center, Durham, NC 27710, USA;3. Molecular Physiology Institute, Duke University Medical Center, Durham, NC 27710, USA;4. Department of Cellular Biology and Anatomy, Georgia Regents University, Augusta, GA 30912;5. Department of Surgery, Division of Cardiothoracic Surgery, Duke University Medical Center, Durham, NC 27710, USA;1. Department of Pharmacology, School of Pharmacy, Nantong University, Qi Xiou Road 19, Nantong 226001, Jiangsu, China;2. Department of Biochemistry & Molecular Biology, Medical College, Nantong University, Qi Xiou Road 19, Nantong, 226001, Jiangsu, China |
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| Abstract: | This paper demonstrates that the mitochondrial isoenzyme of creatine kinase (CKm) can be solubilized from rabbit heart mitochondria, the outer membrane of which has been removed or at least broken by a digitonin treatment or a short hypotonic exposure, but which has retained an important part of the capacity to phosphorylate ADP. Phosphate, ADP, or ATP, at concentrations which are used to study oxidative phosphorylation and creatine phosphate synthesis, solubilize CKm; the same is true with MgCl2 and KCl. The effect of adenine nucleotides does not seem to be due to their interaction with the adenine nucleotide translocase. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis shows that CKm is the main protein released in the described conditions; however, it does not amount to more than 1% of the total protein content of the mitoplasts. When the apparent Km for ATP of CKm was estimated by measuring creatine phosphate synthesis, the values obtained using water-treated mitochondria (0.21 mM) were slightly higher than those of intact mitochondria (0.12 mM) but the difference was not significant. In the former preparation 77% of CKm was in a soluble state. If we can extrapolate these results to intact mitochondria and suppose that in this case a fraction of CKm is also soluble in the intermembrane space, this does not support the theory of functional association between CKm and the adenine nucleotide translocase. |
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