Structural stability of chloroplast coupling factor 1 determined by differential scanning calorimetry and cold inactivation.
At least part of the gamma subunit of the catalytic portion of the chloroplast ATP synthase (CF1) is present in the middle of the alpha3beta3 heterohexamer. Interactions of the alpha/beta subunits with the gamma subunit stabilize the hexameric structure. Surprisingly, neither reduction of the gamma disulfide nor selective proteolysis of alpha, beta and gamma affects the thermal stability of EDTA-treated CF1 preparations, as determined by differential scanning calorimetry. Dissociation of the enzyme in the cold may be monitored by loss of the ATPase activity of CF1 subunit depleted of its epsilon subunit [CF1(-epsilon)]. The rate of cold inactivation of ATPase activity of reduced and alkylated CF1(-epsilon) treated with trypsin in solution was much faster than that CF1(-epsilon)(8.1 versus 38.7 min, respectively, for 50% loss of activity). The increased cold liability of the trypsin-treated enzyme was not a consequence of the cleavage of the gamma. CF1 incubated with trypsin under conditions in which gamma is not cleaved was as cold labile as CF1 with cleaved gamma. Instead, loss of the delta subunit and a few residues from the C-terminal of the beta subunits were responsible for the increased cold liability of the enzyme.
Hightower
K E
KE
Department of Biology, The Johns Hopkins University, Baltimore, Maryland 21218, USA.
McCarty
R E
RE
D020418:Algal Proteins; D002152:Calorimetry, Differential Scanning; D002384:Catalysis; D060365:Chloroplast Proteins; D002736:Chloroplasts / Q000201:enzymology / Q000378:metabolism*; D003080:Cold Temperature; D004795:Enzyme Stability; D007700:Kinetics; D021382:Protein Sorting Signals / Q000378:metabolism*; D006180:Proton-Translocating ATPases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D018724:Spinacia oleracea / Q000201:enzymology / Q000378:metabolism
D020418:Algal Proteins; D060365:Chloroplast Proteins; D021382:Protein Sorting Signals; D006180:Proton-Translocating ATPases