Open AccessInvestigation of Soluble Mitochondrial ATPase by the Reacting Enzyme Sedimentation Method
Author(s) -
CHERNYAK Victor Ya.,
KOZHANOVA Zinayda E.,
CHERNYAK Boris V.,
KOZLOV Igor A.
Publication year - 1979
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1979.tb13220.x
Subject(s) - ultracentrifuge , chemistry , substrate (aquarium) , centrifugation , atpase , enzyme , atp hydrolysis , chromatography , hydrolysis , biochemistry , sedimentation coefficient , mitochondrion , biophysics , biology , ecology
Soluble mitochondrial ATPase from bovine heart (factor F 1 ) loses its activity during ATP hydrolyses. The inactivation is accelerated by moderate pressure, which is generated in an ultracentrifuge cell. The rate of inactivation slows down if the concentration of the substrate (MgATP) is diminished. ATP hydrolysis proceeds at an almost constant rate if the substrate concentration is as low as 0.05 mM. One intersubunit cross‐link formed by dimethylsuberimidate per molecule of factor F 1 , prevents its inactivation during the ATPase reaction both without pressure and in an ultracentrifuge. Sedimentation coefficients measured by the reacting enzyme centrifugation method of both unmodified factor F 1 at a low (about 0.05 mM MgATP) substrate concentration and of its dimethylsuberimidate cross‐linked form in the presence of 10 mM MgATP, were determined to be s 20,w = 12.4 ± 0.4 S. The value is the same as that obtained by the conventional boundary sedimentation method in the absence of the substrate. This result testifies to the fact that the conformation of reacting factor F 1 in solution is similar to that of the enzyme in the absence of the substrate.