Characterization of sulfhydryl groups on chloroplast coupling factor 1 exposed by heat activation

Biochemistry
L C Cantley, G G Hammes

Abstract

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.

Citations

Mar 1, 1985·Photosynthesis Research·S Merchant, B R Selman
Apr 1, 1981·Mutation Research·J O Nelson, P F Warren
Jul 3, 1979·Biochimica Et Biophysica Acta·B A Baird, G G Hammes
Oct 1, 1982·Proceedings of the National Academy of Sciences of the United States of America·L M AmzelP L Pedersen
Jun 1, 1994·Phytochemistry·M J IraburuE Santiago
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Nov 20, 1984·Biochemistry·B Snyder, G G Hammes
Apr 23, 1985·Biochemistry·B Snyder, G G Hammes
Nov 13, 2019·Methods and Applications in Fluorescence·Herbert Schneckenburger
Nov 30, 1976·Biochimica Et Biophysica Acta·N Nelson

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