Adamski J, Husen B, Thole H H, Groeschel-Stewart U, Jungblut P W
Max-Planck-Institut für experimentelle Endokrinologie, Hannover, Germany.
Biochem J. 1993 Dec 15;296 ( Pt 3)(Pt 3):797-802. doi: 10.1042/bj2960797.
We report on the discovery of interactions of porcine endometrial 17 beta-oestradiol dehydrogenase with actin. The 17 beta-oestradiol dehydrogenase of porcine uteri is an essentially unidirectional enzyme compounded in specialized organelles. The enzyme activity in Brij 35 extracts of the particulate fraction of epithelial cells sedimenting between 1800 and 11,000 g(av). was collected by immunoadsorption and eluted at low pH. The eluate contained three proteins of 32, 45 and 80 kDa as shown by SDS/PAGE and silver staining. They were identified by amino acid sequencing and immunotyping as oestradiol dehydrogenase (32 kDa), actin (45 kDa) and a covalent dehydrogenase-actin complex (80 kDa). Disulphides, aldimines, periodate-degradable bonds and hydrophobic interactions were excluded as linkages in the 80 kDa protein. The epsilon-(gamma-glutamyl)-lysine nature of the covalent cross-link was recognized by narrow-bore h.p.l.c. analysis of enzymic digests of electro-eluted 80 kDa material. An involvement of the actin anchor in positioning of the oestradiol dehydrogenase-containing organelles according to metabolic requirements is discussed.
我们报告了猪子宫内膜17β-雌二醇脱氢酶与肌动蛋白相互作用的发现。猪子宫的17β-雌二醇脱氢酶是一种主要存在于特殊细胞器中的单向酶复合物。在上清液中，介于1800至11,000g（平均）之间沉淀的上皮细胞颗粒部分的Brij 35提取物中的酶活性，通过免疫吸附收集，并在低pH下洗脱。如SDS / PAGE和银染所示，洗脱液包含三种蛋白质，分子量分别为32、45和80 kDa。通过氨基酸测序和免疫分型，它们被鉴定为雌二醇脱氢酶（32 kDa）、肌动蛋白（45 kDa）和一种共价脱氢酶-肌动蛋白复合物（80 kDa）。二硫键、醛亚胺、高碘酸盐可降解键和疏水相互作用被排除为80 kDa蛋白质中的连接方式。通过对电洗脱的80 kDa物质的酶解产物进行窄孔高效液相色谱分析，确定了共价交联的ε-（γ-谷氨酰基）-赖氨酸性质。讨论了肌动蛋白锚在根据代谢需求定位含雌二醇脱氢酶的细胞器中的作用。
