Purification of active recombinant trypanosome alternative oxidase.
Trypanosome alternative oxidase (TAO) is the terminal oxidase of the respiratory chain in long slender bloodstream forms of African trypanosomes. TAO is a cytochrome-independent, cyanide-insensitive quinol oxidase. These characteristics are distinct from those of the bacterial quinol oxidases, proteins that belong to the heme-copper terminal oxidase superfamily. The inability to purify stable TAO has severely hampered biochemical studies of the alternative oxidase family. In the present study, we were able to purify recombinant TAO to homogeneity from Escherichia coli membranes using the detergent digitonin. Kinetic analysis of the purified TAO revealed that the specific inhibitor ascofuranone is a competitive inhibitor of ubiquinol oxidase activity.
Nihei
Coichi
C
Department of Biomedical Chemistry, Graduate School of Medicine, University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan.
Fukai
Yoshihisa
Y
Kawai
Keisuke
K
Osanai
Arihiro
A
Yabu
Yoshisada
Y
Suzuki
Takashi
T
Ohta
Nobuo
N
Minagawa
Nobuko
N
Nagai
Kazuo
K
Kita
Kiyoshi
K
D000818:Animals; D004591:Electrophoresis, Polyacrylamide Gel; D004791:Enzyme Inhibitors / Q000494:pharmacology; D024101:Mitochondrial Proteins; D010088:Oxidoreductases / Q000037:antagonists & inhibitors / Q000302:isolation & purification*; D010940:Plant Proteins; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors / Q000302:isolation & purification; D012717:Sesquiterpenes / Q000494:pharmacology; D014345:Trypanosoma / Q000201:enzymology*; D000099188:Alternative Oxidase
D004791:Enzyme Inhibitors; D024101:Mitochondrial Proteins; D010088:Oxidoreductases; D010940:Plant Proteins; D011994:Recombinant Proteins; D012717:Sesquiterpenes; D000099188:Alternative Oxidase; C006640:ascofuranone