Effect of proteolytic digestion on the Ca2+-ATPase activity and subunits of latent and thiol-activated chloroplast coupling factor 1.
The activation by proteases of the Ca2+-dependent ATPase of chloroplast coupling factor 1 (CF1) has been investigated. Using low concentrations of papain and trypsin, the increase in ATPase activity and the degradation of the five subunits of CF1 were compared. Sodium dodecyl sulfate-gel electrophoresis of protease-treated CF1 revealed that the delta subunit was very rapidly degraded and that the alpha and beta subunits were clipped. The gamma and epsilon subunits were more resistant to digestion. The modification of the alpha subunit of latent CF1 most closely correlated with the activation of Ca2+-ATPase activity. Trypsin treatment of dithiothreitol-activated CF1 resulted in a very rapid increase in Ca2+-ATPase activity and a corresponding rapid cleavage of the gamma subunit to a 25,000-dalton species. With more prolonged treatment, the 25,000-dalton species was cleaved to fragments of 14,000 and 11,000-daltons. Dithiothreitol treatment did not alter the rate of attack on the other subunits. The gamma subunit of heat-activated CF1 was also more susceptible to protease digestion. The increased protease sensitivity of the gamma subunit of soluble CF1 after treatment with dithiothreitol or heat mimics the increased protease sensitivity of the gamma subunit of bound CF1 when thylakoids are treated with trypsin during illumination (Moroney, J. V., and McCarty, R. E. (1982) J. Biol. Chem. 257, 5915-5920). These results suggest that the conformational changes that occur when purified CF1 is exposed to dithiothreitol are similar to those that CF1 bound to thylakoid membranes undergoes under illumination.
D000251:Adenosine Triphosphatases / Q000378:metabolism*; D000252:Calcium-Transporting ATPases / Q000378:metabolism*; D002736:Chloroplasts / Q000201:enzymology*; D004789:Enzyme Activation; D007700:Kinetics; D046911:Macromolecular Substances; D010086:Oxidative Phosphorylation Coupling Factors / Q000378:metabolism*; D010206:Papain / Q000494:pharmacology*; D010944:Plants / Q000201:enzymology; D006180:Proton-Translocating ATPases; D014357:Trypsin / Q000494:pharmacology*
D046911:Macromolecular Substances; D010086:Oxidative Phosphorylation Coupling Factors; D014357:Trypsin; D010206:Papain; D000251:Adenosine Triphosphatases; D006180:Proton-Translocating ATPases; D000252:Calcium-Transporting ATPases