Fedosov S N, Belousova L V
Biokhimiia. 1989 Jan;54(1):54-67.
It was found that in the octameric form of mitochondrial creatine kinase (Mr = 340 kD), only 52% of active centers bind Mg-ADP into a E-Mg-ADP-creatine complex with the dissociation constant, K(Cr)ADP, of 0.105 mM, which is close to the Km value for the enzyme (0.072 mM). In the dimeric form of cytoplasmic creatine kinase (Mr = 82 kD), 100% of active centers bind Mg--ADP; the K(Cr)ADP value (0.11 mM) is close to the Km value for the given enzyme preparation (0.083 mM). All active centers of rabbit muscle cytoplasmic creatine kinase were shown to form an analog of the transition state complex (ATSC) - E-Mg-ADP-NO3- -creatine. The constant for Mg-ADP dissociation from ATSC is identical for all centers of cytoplasmic creatine kinase and equals to 6.0 microM. The curves for ATSC saturation with Mg-ADP in the presence of iodacetamide for mitochondrial creatine kinase were constructed and computer analyzed. It was shown that in the octameric form of the enzyme only 54 +/- 13% of subunits can form ATSC. The constant for Mg-ADP dissociation from ATSC, KATSCADP is equal to 1.9 +/- 0.8 microM. It was concluded that 50% of subunits of the octameric form of mitochondrial creatine kinase are not involved in the catalytic act due to masking of their active centres and their inability to form transition state complexes. A model of regulation of cell supply with high energy compounds, e.g., ATP, creatine phosphate, via association-dissociation of mitochondrial creatine kinase oligomers is proposed.
研究发现，在线粒体肌酸激酶的八聚体形式（Mr = 340 kD）中，只有52%的活性中心能将Mg-ADP结合形成E-Mg-ADP-肌酸复合物，其解离常数K(Cr)ADP为0.105 mM，这与该酶的Km值（0.072 mM）相近。在细胞质肌酸激酶的二聚体形式（Mr = 82 kD）中，100%的活性中心能结合Mg-ADP；K(Cr)ADP值（0.11 mM）与给定酶制剂的Km值（0.083 mM）相近。已证明兔肌肉细胞质肌酸激酶的所有活性中心都能形成过渡态复合物类似物（ATSC）——E-Mg-ADP-NO3- -肌酸。Mg-ADP从ATSC解离的常数在细胞质肌酸激酶的所有中心都相同，等于6.0 microM。构建了在线粒体肌酸激酶存在碘乙酰胺时Mg-ADP对ATSC饱和的曲线并进行了计算机分析。结果表明，在该酶的八聚体形式中，只有54±13%的亚基能形成ATSC。Mg-ADP从ATSC解离的常数KATSCADP等于1.9±0.8 microM。得出的结论是，线粒体肌酸激酶八聚体形式中50%的亚基由于其活性中心被掩盖且无法形成过渡态复合物，不参与催化作用。提出了一种通过线粒体肌酸激酶寡聚体的缔合-解离来调节细胞高能化合物（如ATP、磷酸肌酸）供应的模型。
