KINETICS AND THIOREDOXIN SPECIFICITY OF THIOL MODULATION OF THE CHLOROPLAST H-ATPASE()

The kinetics of thiol modulation of the chloroplast H+-ATPase (CF0CF1) in membrana were analyzed by employing thioredoxins that were kept re duced by 0.1 mM dithiothreitol. The kinetics of thiol modulation depen d on the extent of the proton gradient. The process is an exponential function of the thioredoxin concentration and reaction time and can be described by an irreversible second order reaction. The results indic ate that the formation of the complex between thioredoxin and CF0CF1 i s slow compared with the subsequent reduction step. Furthermore we hav e compared the efficiencies of the Escherichia coli thioredoxin Trx an d the two chloroplast thioredoxins Tr-m and Tr-f. The second order rat e constants are 0.057 (Tr-f), 0.024 (Trx), and 0.010 s(-1) mu M-1 (Tr- m) suggesting that Tr-f rather than Tr-m is the physiological reductan t for the chloroplast ATPase. The often employed artificial reductant dithiothreitol exhibits a second order rate constant in thiol modulati on of 1.02.10(-6) s(-1) mu M-1.