KINETICS, REGULATION AND MECHANISMS OF BIOCHEMICAL REACTIONS

(1)Mg（II）-ATP依赖性磷酸蛋白由两种类型的 亚单位。 SDS-凝胶过程中催化亚基C在38 KDa处发生迁移 电泳和调节亚基M在31 K Da。这种酶是 不活跃，孤立。 它被蛋白激酶FA激活， 与糖原合成酶激酶-3和MgATP相同。 该酶的定量 在粗提物中的含量表明它以高浓度存在（约50 nM）。 此外，纯化的酶的最大比活性为 酶浓度依赖性。 进一步的研究表明，这种酶 由两个调节剂和一个催化亚基组成， 从（i）凝胶过滤和蔗糖梯度离心获得的数据 实验，（ii）银染色SDS-聚丙烯酰胺的密度扫描 凝胶，（iii）基于分离的氨基酸分析的计算 （iv）变性亚基的UV光谱的去卷积分析; 酶复合物，和（v）酶的稳态动力学分析 通过添加调节剂亚基抑制。 一种调节剂与 催化亚单位，而另一个是可解离的，并作为一个 底物的竞争性抑制剂。 因此，它提供了一种机制， 保护以低浓度存在的磷蛋白。 (2)跨膜蛋白相互作用模型的建立 和膜电位。 该理论可以用来提供合理的 解释能量转换和解释两个先验 独立的反应可以通过酶偶联。 (3)细胞内钙和pH浓度的快速变化， 紫球海胆精子与精子作用 鸡蛋的大小被测量。

(1) The Mg(II)-ATP-dependent protein phosphate is composed of two types of subunits. The catalytic subunit C migrates at 38K Da during SDS-gel electrophoresis and the modulator subunit M at 31K Da. This enzyme is inactive as isolated. It is activated by protein kinase FA which is the same as glycogen synthase kinase-3, and MgATP. Quantitation of this enzyme in crude extracts shows that it is present in high concentration (about 50 nM). In addition, the maximal specific activity of the purified enzyme is enzyme concentration dependent. Further study shows that the enzyme is composed of two modulator and one catalytic subunit as evidenced by the data obtained from (i) gel filtration and sucrose gradient centrifugation experiments, (ii) densitometric scans of silver stained SDS-polyacrylamide gels, (iii) calculations based on amino acid analysis of the separated subunits; (iv) deconvolution analysis of the UV spectra of the denatured enzyme complex, and (v) steady-state kinetic analysis of the enzyme inhibition by added modulator subunit. One modulator binds very tightly to the catalytic subunit, while the other is dissociable and functions as a competitive inhibitor for the substrate. Thus, it provides a mechanism for protecting the phosphoproteins present in low concentrations. (2) Development of a model for interactions between transmembrane proteins and the membrane potential. The theory can be used to provide a reasonable interpretation for energy transduction and for explaining how two a priori independent reactions can be coupled by an enzyme. (3) Rapid intracellular changes in both calcium and pH concentration in sperm of sea urchin Strongylocentratus purpuratus caused by speract and by egg coat were measured.