Characterization of sulfhydryl groups on chloroplast coupling factor 1 exposed by heat activation.
Two sulfhydryl groups in the gamma subunit of solubilized chloroplast coupling factor 1 (CF1) are exposed by heat activating the enzyme. These two groups have been selectively labeled with [3H]-N-ethylmaleimide, N-[p-(2-benzoxazolyl)phenyl]maleimide (NBPM), and N-(4-dimethylamino-3,5-dinitrophenyl)maleimide (DDPM). Modifying these groups did not appreciably effect Ca2+-ATPase activity, the ability of the enzyme to bind quercetin and 1,N6-ethenoadenosine diphosphate (epsilonADP), or the ability of the enzyme to react with 7-chloro-4-nitrobenzo-2-oxa-1,3-diazole (NBD-C1). Fluorescence resonance energy transfer was used to measure the distance from the sulfhydryl groups on the gamma subunit to the quercetin sites, the NBD-C1 reactive sites, and the tight nucleotide sites on CF1 using the donor-acceptor pairs NBPM-quercetin, NBPM-NBD, and epsilonADP-DDPM, respectively. The distance from the sulfhydryl groups to the quercetin sites was found to be less than 30 A, the distance to the NBD-C1 reactive was 34-41 A, and the distance to the tight nucleotide sites as greater than 40 A. A three-dimensional symmetrical model is proposed for the relative positions of sites on CF1.
D000251:Adenosine Triphosphatases / Q000378:metabolism; D001665:Binding Sites; D002118:Calcium / Q000494:pharmacology; D002736:Chloroplasts / Q000378:metabolism*; D004735:Energy Transfer; D004789:Enzyme Activation; D005033:Ethylmaleimide; D006358:Hot Temperature; D008301:Maleimides; D008433:Mathematics; D010940:Plant Proteins / Q000032:analysis* / Q000378:metabolism; D010944:Plants; D011485:Protein Binding; D011487:Protein Conformation; D011955:Receptors, Drug; D013050:Spectrometry, Fluorescence; D013438:Sulfhydryl Compounds / Q000032:analysis
D008301:Maleimides; D010940:Plant Proteins; D011955:Receptors, Drug; D013438:Sulfhydryl Compounds; D000251:Adenosine Triphosphatases; D005033:Ethylmaleimide; D002118:Calcium