THE MITOCHONDRIAL ATP SYNTHASE OF TRYPANOSOMA-BRUCEI - ISOLATION AND CHARACTERIZATION OF THE INTACT F1 MOIETY

作     者：WILLIAMS, N FRANK, PH 

作者机构：Uniformed Services University of the Health Sciences Department of Biochemistry Bethesda MD 20814-4799. 

出 版 物：《MOLECULAR AND BIOCHEMICAL PARASITOLOGY》 (分子与生化寄生虫学)

年 卷 期：1990年第43卷第1期

页      面：125-132页

核心收录：

学科分类：0710[理学-生物学] 071010[理学-生物化学与分子生物学] 1001[医学-基础医学(可授医学、理学学位)] 07[理学] 

主　　题：Bioenergetics Trypanosoma brucei F 1 -ATPase Purification Reconstitution Kinetics PEP phosphoenolpyruvate LDH lactate dehydrogenase PK pyruvate kinase BSA bovine serum albumin SDS sodium dodecyl sulfate TEMED N , N , N ′, N ′-tetramethylenediamine 

摘      要：This report describes the first isolation and molecular characterisation of the mitochondrial F1-ATPase from Trypanosoma brucei. The isolation procedure utilized is a modified chloroform extraction procedure. In contrast to earlier reports on the F1-ATPase from other trypanosomatids, the F1-ATPase we have isolated from the procyclic form of T. brucei a complex composed of five distinct subunits. Apparent molecular weights of these subunits are 55000[.alpha.], 42000 [.beta.], 32000 [.gamma.), 22000 [.delta.], and 17000 [.epsilon.]. The F1 moiety which possesses the active site of the H+-ATPase has an ATPase activity in the standard Tris-HCl coupled enzyme assay with Vmax of 22.96 .*** min-1 (mg protein)-1 and a Km value of 0.60 mM. This ATPase activity is cold labile and is not susceptible to oligomycin inhibition as is the membrane bound enzyme. Upon reconstitution with F1-ATPase depleted membranes (urea particles) the ATPase regains oligomycin sensitivity to the same extent as that found in the intact inner membrane vesicles. ATP synthesis is also restored to these particles upon reconstitution with F1. These results indicate that this F1-ATPase as isolated is intact with respect to all the critical H+-ATPase functions.