Open AccessPurification of a cytochrome aa 3 terminal oxidase from protoplast membrane vesicles of Micrococcus luteus
Author(s) -
Heinz Gerd,
Dose Klaus,
Nawroth Thomas
Publication year - 1994
Publication title -
fems microbiology letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.899
H-Index - 151
eISSN - 1574-6968
pISSN - 0378-1097
DOI - 10.1111/j.1574-6968.1994.tb07281.x
Subject(s) - micrococcus luteus , cytochrome c oxidase , cytochrome , protoplast , biochemistry , heme , vesicle , oxidase test , electron transport complex iv , chromatography , antimycin a , chemistry , biology , enzyme , electron transport chain , membrane , escherichia coli , gene
A cytochrome aa 3 terminal oxidase was isolated from protoplast membrane vesicles of Micrococcus luteus grown under aerobic conditions. The purified complex showed similarities to cytochrome c oxidase (EC 1.9.3.1) of the electron transport chain of mitochondria and many prokaryotes. The enzyme was solubilized by subsequent treatment with the detergents CHAPS and n‐dodecyl‐β‐d‐maltoside and purified by ion‐exchange chromatography using poly‐L‐lysine agarose and TMAE‐fractogel‐650 (S) columns, followed by hydroxyapatite chromatography. The purified complex is composed of two major subunits with apparent molecular masses of 54 and 32 kDa. After purification the isolated enzyme contains 12.1 nmol of heme A (mg protein) −1 and exhibits absorption maxima at 424 nm and 598 nm in the oxidized state and at 442 nm and 599 nm in the reduced state. The CO‐difference spectrum shows peaks at 428 and 590 nm which is indicative of heme a 3 , furthermore oxygen consumption was found to be sensitive to cyanide.