Trofimova M E, Moiseeva N S
Department of Biochemistry, School of Biology, Moscow State University, USSR.
Biochem Int. 1989 Sep;19(3):603-13.
It has been found that at pH 7.4 and 2 mg/ml protein, bovine heart mitochondrial creatine kinase (CKm) contains less than 10% of the dimer. The constant for the CKm octamer dissociation into dimers, Kd, in the presence of substrates forming an analog of the complex of the transient state was found to be equal to 4.9 10(-17) M3. Using this value, the experimental conditions were chosen so as to achieve a practically complete dissociation of the octamer into dimers. Evidence has been obtained suggesting that the octamer does not dissociate into dimers during the time course of the kinetic experiments; the corresponding kinetic parameters of the CKm octamer and dimer are as follows: KMgATPm = 82 microM and 42 microM; KCrm = 8.1 mM and 3.4 mM; Vf = 61 and 60 micrograms-equiv. H+ min-1 mg-1; KMgADPm = 43 microM and 17 microM, KCPm = 0.68 mM and 0.23 mM; Vr = 162 and 111 micrograms-equiv. H+ min-1 mg-1. The experimental and calculated data shed additional light on the physiological role of CKm.
已发现，在pH 7.4和蛋白质浓度为2 mg/ml的条件下，牛心线粒体肌酸激酶（CKm）的二聚体含量不到10%。在存在形成瞬态复合物类似物的底物时，CKm八聚体解离为二聚体的常数Kd被发现等于4.9×10⁻¹⁷ M³。利用该值，选择了实验条件以实现八聚体几乎完全解离为二聚体。已获得的证据表明，在动力学实验过程中八聚体不会解离为二聚体；CKm八聚体和二聚体的相应动力学参数如下：KMgATPm = 82 μM和42 μM；KCrm = 8.1 mM和3.4 mM；Vf = 61和60微克当量H⁺ min⁻¹ mg⁻¹；KMgADPm = 43 μM和17 μM，KCPm = 0.68 mM和0.23 mM；Vr = 162和111微克当量H⁺ min⁻¹ mg⁻¹。实验和计算数据为CKm的生理作用提供了更多线索。
