Expression,purification and characterization of the soluble Cu_A domain of cytochrome c oxidase of Paracoccus versutus

作     者：LI Lianzhi SONG Aixin XIE Yi HUANG Zhongxian Ellen de Waal Urszula Kolczak Gerard W.Canters 

作者机构：Chemical Biology LaboratoryDepartment of ChemistryFudan UniversityShanghai 200433China State Key Laboratory of GeneticsSchool of Life ScienceFudan UniversityShanghai 200433China Leiden Institute of ChemistryLeiden UniversityPO Box 95022300 RA LeidenThe Netherlands 

出 版 物：《Chinese Science Bulletin》 (CHINESE SCIENCE BULLETIN)

年 卷 期：2001年第46卷第19期

页      面：1608-1612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] 081704[工学-应用化学] 07[理学] 08[工学] 0817[工学-化学工程与技术] 

基　　金：This work was supported by the National Natural Science Foundation of China (Grant No. 39990600) 

主　　题：cytochrome coxidase CuA domain protein gene expression purification spectra. 

摘      要：The key subunit II of cytochrome c oxidase (CcO) contains a soluble binuclear copper center (CuA) domain. The CUA domain of Paracoccus versutus was cloned, expressed, purified and characterized. The gene encoding the CUA domain in pETlld vector was expressed in E. coli BL21 (DE3). The results showed that the CuA domain was expressed mostly in inclusion bodies and the CUA domain protein synthesized in E. coli cells represents approximately 10 percent of the total cellular proteins. Dissolved in urea, dia-lyzed and recombined with Cu+/Cu2+ and purified by the Q-sepharose fast flow anion-exchange column and Sephadex G-75 gel filtration column, the soluble purple-colored protein, which shows a single band in electrophoresis, was obtained. The UV-visible absorption spectrum of CUA domain showed that there are intense band at 478 nm and a shoulder peak at 530 nm, and two weak bands at 360 and 806 nm respectively, which can be assigned to the charge transfer and the interactions of obitals of Cu-S and Cu-Cu