Modifications of the gamma subunit of chloroplast coupling factor 1 alter interactions with the inhibitory epsilon subunit.
The treatment of chloroplast coupling factor 1 (CF1) with dithiothreitol or with trypsin modifies the gamma subunit. Reduction of the gamma subunit disulfide bond in CF1 in solution with dithiothreitol enhances the dissociation of epsilon (Duhe, R. J., and Selman, B. R. (1990) Biochim. Biophys. Acta 1017, 70-78). The Ca(2+)-ATPase activity of either oxidized or reduced CF1 increases as the enzyme is diluted. Added epsilon subunit inhibits the Ca(2+)-ATPase activity of both forms of the diluted CF1, suggesting that epsilon dissociation is the cause of activation by dilution. Half-maximal activation occurred at much higher concentrations of the reduced CF1, indicating that reduction decreases the affinity for epsilon about 20-fold. Immunoblotting techniques show that there is only one epsilon subunit/CF1 in intact chloroplasts, in thylakoid membranes, and in solution. No epsilon is released from CF1 in thylakoids under conditions of ATP synthesis. The gamma subunit of CF1 in illuminated thylakoids is specifically cleaved by trypsin. CF1 purified from thylakoids treated with trypsin in the light is deficient in epsilon subunit, and has a high rate of ATP hydrolysis. Added epsilon neither inhibits the ATPase activity of, nor binds tightly to the cleaved enzyme.
Soteropoulos
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Section of Biochemistry, Molecular and Cell Biology, Cornell University, Ithaca, New York 14853.
Süss
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McCarty
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D000255:Adenosine Triphosphate / Q000096:biosynthesis / Q000378:metabolism; D015153:Blotting, Western; D000252:Calcium-Transporting ATPases / Q000037:antagonists & inhibitors / Q000378:metabolism; D002736:Chloroplasts / Q000201:enzymology; D004229:Dithiothreitol / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004789:Enzyme Activation; D006868:Hydrolysis; D007425:Intracellular Membranes / Q000201:enzymology; D009995:Osmosis; D010084:Oxidation-Reduction; D006180:Proton-Translocating ATPases / Q000378:metabolism*; D014357:Trypsin / Q000378:metabolism
D000255:Adenosine Triphosphate; D014357:Trypsin; D006180:Proton-Translocating ATPases; D000252:Calcium-Transporting ATPases; D004229:Dithiothreitol