Kerr M A
Biochem J. 1980 Jul 1;189(1):173-81. doi: 10.1042/bj1890173.
The assembly of the classical pathway C3 convertase in the fluid phase has been studied. The enzyme is assembled from C2 and C4 on cleavage of these proteins by C1s. Once assembled, the enzyme activity decays rapidly. Kinetic evidence has been obtained that this decay is even more rapid than previously suggested (kdecay is 2.0 min-1 at 37 degrees C). As a result, optimal C3 convertase activity is only observed with high C1s levels, which result in rapid rates of cleavage of C2 and increased rates of formation of the C3 convertase. Using high concentrations of C1s at lower temperatures (22 degrees C) in the presence of excess substrate we have demonstrated kinetically that the enzyme comprises an equimolar complex of C4b and cleaved C2. We have obtained direct evidence from gel-filtration experiments for the role of C2a as the catalytic subunit of the enzyme. C2b appears to mediate the interaction between C4 (or C4b) and C2 at pH 8.5 and at low ionic strength where the interactions can easily be detected. It may therefore be important in the assembly of the enzyme, though it is not involved in the catalytic activity. The decay of the C3 convertase reflects the release of C2a from the C4b x (C2b) x C2a complex, and the stabilizing effect of iodine on the C3 convertase is therefore apparently one of stabilizing the C4b-C2z interaction, which is otherwise weak. C1s is not a part of the C3 convertase enzyme.
已对液相中经典途径C3转化酶的组装进行了研究。该酶由C2和C4在被C1s裂解后组装而成。一旦组装完成，酶活性会迅速衰减。动力学证据表明，这种衰减比之前认为的还要快（在37℃时k衰变率为2.0分钟⁻¹）。因此，只有在高C1s水平下才能观察到最佳的C3转化酶活性，这会导致C2的裂解速度加快以及C3转化酶形成速度增加。在存在过量底物的情况下，于较低温度（22℃）使用高浓度的C1s，我们从动力学上证明该酶由C4b和裂解后的C2的等摩尔复合物组成。我们通过凝胶过滤实验获得了直接证据，证明C2a作为该酶的催化亚基的作用。在pH 8.5和低离子强度下，C2b似乎介导了C4（或C4b）与C2之间的相互作用，在这种情况下这种相互作用很容易被检测到。因此，它在酶的组装中可能很重要，尽管它不参与催化活性。C3转化酶的衰减反映了C2a从C4b x（C2b）x C2a复合物中的释放，因此碘对C3转化酶的稳定作用显然是稳定C4b - C2z相互作用，否则这种相互作用很弱。C1s不是C3转化酶的一部分。
