Study of the structure of the thylakoid membrane-bound chloroplast coupling factor CF1.
The structure of thylakoid membrane-bound chloroplast coupling factor CF1 was studied by limited proteolysis followed by sodium dodecylsulfate polyacrylamide gel electrophoresis and N-terminal sequence analysis. The N-terminal fragment of the alpha-subunit was shown to have an exposed area including the peptide bond R21-E22. The cleavage of this peptide bond caused the alphaK24-V25 bond to be exposed to the outside. In the N-terminal fragment of the beta-subunit, the L14-E15 bond was identified and found to be subject to trypsinolysis. Also, the alphaR140-S141, alphaG160-R161, and betaG102-G103 bonds were accessible to the proteolytic attack. In general, the beta-subunit of membrane-bound CF1 is more sensitive to proteolysis than that of solubilized CF1. The products of proteolysis of the alpha-subunit did not contain the polypeptides typical of the reaction of cleavage of the alphaE17-G18 and alphaE22-V23 bonds in isolated CF1. These results suggest a significant structural difference between soluble and membrane-bound CF1. A number of peptide bonds, alphaG160-R161 in particular, were shown to be shielded from proteolytic attack by papain in illuminated thylakoid membranes, probably as a result of membrane energization. In contrast, the light-induced reduction of the gamma-subunit caused an increase in the accessibility of some peptide bonds to this protease, including the alphaG160-R161 bond.
Malyan
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AN
Institute of Basic Biological Problems, Russian Academy of Sciences, Pushchino, Moscow Region, 142292, Russia. Malyan@issp.serpukhov.su
Vitseva
O I
OI
Strotmann
H
H
D000595:Amino Acid Sequence; D003198:Computer Simulation; D007425:Intracellular Membranes / Q000201:enzymology; D046911:Macromolecular Substances; D008958:Models, Molecular; D010446:Peptide Fragments / Q000737:chemistry; D011487:Protein Conformation; D006180:Proton-Translocating ATPases / Q000737:chemistry* / Q000302:isolation & purification; D018724:Spinacia oleracea / Q000201:enzymology; D020524:Thylakoids / Q000201:enzymology*
D046911:Macromolecular Substances; D010446:Peptide Fragments; D006180:Proton-Translocating ATPases