Activation of Escherichia coli F1-ATPase by lauryldimethylamine oxide and ethylene glycol: relationship of ATPase activity to the interaction of the epsilon and beta subunits.
The stimulation of the ATPase activity of Escherichia coli F1-ATPase by the detergent lauryldimethylamine oxide (LDAO) and the relationship of this activation to removal of the inhibitory epsilon subunit were studied. The detergent caused a dramatic decrease in the affinity of epsilon-depleted enzyme for epsilon subunit, suggesting that release of epsilon is involved in LDAO activation. However, even in the absence of any epsilon subunit, the detergent caused a 140% increase in activity, indicating activation by effects independent of epsilon. In contrast, the addition of 30% ethylene glycol to the reaction buffer caused a modest inhibition of the ATPase activity of epsilon-depleted F1-ATPase but rendered the enzyme insensitive to inhibition by epsilon subunit. This solvent prevented the cross-linking of epsilon to beta by a water-soluble carbodiimide, although epsilon remained linkable to both beta and gamma by dithiobis(succinimidyl propionate). Thus, epsilon was not dissociated from F1-ATPase, but its intimate interaction with the beta subunit was altered. These results suggest that the inhibitory action of epsilon is expressed through its interaction with beta. Kinetic analysis revealed that LDAO activated hydrolysis at both the high- and low-affinity promotional sites, with little change in Km values. Ethylene glycol caused a substantial increase in Km at the low-affinity promotional site and made the enzyme resistant to inhibition by aurovertin D.
Dunn
S D
SD
Department of Biochemistry, University of Western Ontario, London, Canada.
Tozer
R G
RG
Zadorozny
V D
VD
D003432:Cross-Linking Reagents; D003902:Detergents / Q000494:pharmacology*; D004123:Dimethylamines / Q000494:pharmacology*; D004591:Electrophoresis, Polyacrylamide Gel; D004789:Enzyme Activation; D004926:Escherichia coli / Q000201:enzymology*; D019855:Ethylene Glycol; D005026:Ethylene Glycols / Q000494:pharmacology*; D007700:Kinetics; D046911:Macromolecular Substances; D008970:Molecular Weight; D006180:Proton-Translocating ATPases / Q000378:metabolism*; D013501:Surface-Active Agents / Q000494:pharmacology*
D003432:Cross-Linking Reagents; D003902:Detergents; D004123:Dimethylamines; D005026:Ethylene Glycols; D046911:Macromolecular Substances; D013501:Surface-Active Agents; C014518:dodecyldimethylamine oxide; D006180:Proton-Translocating ATPases; D019855:Ethylene Glycol