人骨骼肌磷酸丙糖异构酶多种形式的分离与鉴定

The isolation and characterization of the multiple forms of human skeletal muscle triosephosphate isomerase.

Eber S W, Krietsch W K

Biochim Biophys Acta. 1980 Jul 10;614(1):173-84. doi: 10.1016/0005-2744(80)90178-3.

Abstract

Human skeletal muscle triosephosphate isomeras (D-glyceraldehyde-3-phosphate ketol-isomerase, EC 5.3.1.1) was isolated and resolved by DEAE-cellulose chromatography into three major forms, A, B, and C, which comprise 97% of the total activity. The relative distribution was 25, 46 and 29% respectively. 2. The A and C forms are homodimers, alpha alpha and beta beta, and form B is the heterodimer, alpha beta. Reassociation studies from guanidinium chloride have indicated that A, B, and C are not conformers. Although these studies revealed the existence of two different chains, the amino acid analysis showed no significant variance. Since no differences were obsrved in Ouchterlony and Mancini tests or in immunotitration, the three fors are assumed to be immunologically identical. 3. The three forms have the same specific activity, Michaelis constants, pH optimum, activation energy, inhibition by metabolites and heat stability. Only with increasing ionic strength did the V and Km values differ. 4. The two poypeptide chains (alpha and beta) appear to be identical (amino acid composition, molecular weight and antigenity), and since the electrophoretic banding pattern changed with cell aging, it is concluded that the multiple forms of trisephosphate isomerase are the consequence of minor post-synthetic alteration(s) of form A.

摘要

人骨骼肌磷酸丙糖异构酶（D-甘油醛-3-磷酸酮醇异构酶，EC 5.3.1.1）经DEAE-纤维素色谱分离并解析为三种主要形式，A、B和C，它们占总活性的97%。相对分布分别为25%、46%和29%。2. A和C形式是同二聚体，αα和ββ，而B形式是异二聚体，αβ。来自氯化胍的重缔合研究表明，A、B和C不是构象异构体。尽管这些研究揭示了存在两条不同的链，但氨基酸分析显示没有显著差异。由于在双向免疫扩散和 Mancini 试验或免疫滴定中未观察到差异，因此假定这三种形式在免疫学上是相同的。3. 这三种形式具有相同的比活性、米氏常数、最适pH值、活化能、代谢物抑制作用和热稳定性。只有随着离子强度的增加，V和Km值才会有所不同。4. 两条多肽链（α和β）似乎是相同的（氨基酸组成、分子量和抗原性），并且由于电泳条带模式随细胞老化而变化，因此得出结论，磷酸丙糖异构酶的多种形式是A形式合成后轻微改变的结果。