金属置换碳酸酐酶及其生成动力学

METAL SUBSTITUTED CARBONIC ANHYDRASE AND THEIR FORMATION KINETICS

本文报道了金属置换碳酸酐酶的活性研究。指出了离子半径的大小,配位构型的要求,热力学稳定性和配位体交换动力学是决定各种金属置换碳酸酐酶活性的重要因素。并用金属置换法测定了Ni(Ⅱ)与apo-CA形成金属酶的反应动力学速率常数。在25℃、pH7.47时,K_(fNi(Ⅱ)-CA)为14.01·mol^(-1)sec^(-1),较之于Zn(Ⅱ)-CA的速率常数小10~3倍。Cd(Ⅱ)-CA的生成动力学为一快反应,采用配位竞争法,在邻菲罗啉配位剂存在下,测得了Cd(Ⅱ)-CA生成动力学的条件速率常数在o-phen浓度为1.96和2.92×10^(-4)mol·1^(-1)时分别为137.1和114.61·mol^(-1)sec^(-1)。这一研究提示了镉对生物体的毒性作用的一个重要方面是使锌酶失活。而配位剂的存在大大降低了镉与脱辅基锌酶的反应动力学,这也是它们显示解毒功能所在。

The activities of Co(II ), Mn(II ), Ni(II ), Cu(II ) and Cd(II ) metal substituted carbonic anhydrases(CA) have been studied. The results indicate that the crucial factors manipulating the enzyme activity are the metal ionic radius, the coincidence of coordination geometry, the thermodynamic stability constants of metal ions with apo-CA and the ligand (especially the water molecule) exchange kinetics on metal ions. The formation rate constant of Ni(II )-CA has been spectrophotometrically measured by a metal substitution reaction. Under the conditions of 251 and pH 7.57 the kf, Ni(II)-CA= 14.0 1 mol-1 sec-1 which is about 103 times slower than the formation rate constant of Zn(II )-CA. Cadmium ion forming Cd(II )-CA with apo-CA has rather fast kinetics. Usually, it is difficult to determine the rate constant. Here, the apparent rate constants of Cd(II)-CA formation in the presence of competitive ligand such as o-phenanthroline has been successfully estimated. With different condition of o-phen at 1.96 and 2.92 × 10-4mol·l-1 the rate constants of Cd(II )-CA formation, 137.1 and 114.61·mol-1 sec-1 have been obtained respectively. The studies remind us that the toxicity of cadmium is the results of the inactivation of essential zinc-containing metallocnzym.es. The chelating agents function as a scavenger of toxic material cadmium by inhibiting the formation of Cd (II )-enzyme.