Conformational change in beef-heart mitochondrial F1 ATPase to ATP synthesis mode induced by dimethylsulfoxide and ATP revealed by sulfhydryl group labeling.
Treatment of beef-heart mitochondrial F1 ATPase with 5,5'-dithiobis(2-nitrobenzoic acid) (DTNB) results in the incorporation of 1 mol DTNB/mol F1 without loss of ATPase activity. Incorporation is not prevented by ATP. Labeling occurs predominantly on an alpha-subunit, but also with a significant degree of modification of gamma- and epsilon-subunits. It is suggested that the modified sulfhydryl groups of the alpha-, gamma- and epsilon-subunits are in proximity so that only one can be modified by the reagent. Guanidine hydrochloride (0.3 M) dissociates F1 into its subunits. Eight sulfhydryl groups/mol F1 can be modified under these conditions. Guanidine hydrochloride does not cause dissociation of F1 in the presence of 30% (v/v) dimethylsulfoxide (Me2SO) and 2 mM ATP. Under these conditions a second molecule of DTNB is incorporated into F1 with nearly equal modification of the epsilon-subunit and an alpha-subunit. It is proposed that Me2SO and ATP induce a more stable conformation of F1, which is resistant to dissociation by guanidine hydrochloride, but in which the site of reaction with DTNB is made more accessible by the guanidine hydrochloride to permit the simultaneous modification of an alpha-subunit and the epsilon-subunit. This conformation is probably that which occurs during ATP synthesis by F1 in the presence of Me2SO.
Beharry
S
S
Department of Biochemistry, University of British Columbia, Vancouver, Canada.
Bragg
P D
PD
D000255:Adenosine Triphosphate / Q000494:pharmacology*; D000818:Animals; D002417:Cattle; D003545:Cysteine; D004121:Dimethyl Sulfoxide / Q000494:pharmacology*; D004228:Dithionitrobenzoic Acid / Q000494:pharmacology; D019791:Guanidine; D006146:Guanidines / Q000494:pharmacology; D008929:Mitochondria, Heart / Q000201:enzymology*; D011487:Protein Conformation / Q000187:drug effects; D006180:Proton-Translocating ATPases / Q000378:metabolism*
D006146:Guanidines; D000255:Adenosine Triphosphate; D004228:Dithionitrobenzoic Acid; D006180:Proton-Translocating ATPases; D019791:Guanidine; D003545:Cysteine; D004121:Dimethyl Sulfoxide