Open AccessCross‐reconstitution of the F O F 1 ‐ATP synthases of chloroplasts and Escherichia coli with special emphasis on subunit δ
Author(s) -
ENGELBRECHT Siegfried,
DECKERSHEBESTREIT Gabriele,
ALTENDORF Karlheinz,
JUNGE Wolfgang
Publication year - 1989
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1989.tb14750.x
Subject(s) - photophosphorylation , thylakoid , chloroplast , vesicle , escherichia coli , delta , quenching (fluorescence) , biology , atp synthase , chemiosmosis , electrochemical gradient , biochemistry , biophysics , tricine , membrane , fluorescence , engineering , physics , quantum mechanics , gene , enzyme , aerospace engineering
F O F 1 ‐ATP synthases catalyse ATP formation from ADP and P i by using the free energy supplied by the transmembrane electrochemical potential of the proton. The δ subunit of F 1 plays an important role at the interface between the channel portion F o and the catalytic portion F 1 . In chloroplasts it can plug the protonic conductance of CF o and in Escherichia coli it is required for binding of EF 1 to EF o . We wanted to know whether or not δ of one species was effective between F o and F 1 of the other species and vice versa. To this end the respective coupling membrane (thylakoids, everted vesicles from E . coli) was (partially) depleted of F 1 and purified F 1 , F 1 (‐δ), and δ were added in various combinations to the F 1 ‐depleted membranes. The efficiency or reconstitution was measured in thylakoids via the rate of phenazinemethosulfate‐mediated cyclic photophosphorylation and in E. coli everted vesicles via the degree of 9‐amino‐6‐chloro‐2‐methoxyacridine fluorescence quenching. Addition of CF 1 to partially CF 1 ‐depleted thylakoid vesicles restored photophosphorylation to the highest extent. CF 1 (‐δ)+chloroplast δ, EF 1 , EF 1 (‐δ)+ E. coli δ were also effective but to lesser extent. CF 1 (‐δ)+ E. coli δ and EF 1 (‐δ)+chloroplast δ restored photophosphorylation to a small but still significant extent. With F 1 ‐depleted everted vesicles prepared by repeated EDTA treatment of E. coli membranes, addition of CF 1 , CF 1 (‐δ)+chloroplast δ and CF 1 (‐δ)+ E. coli δ gave approximately half the extent of 9‐amino‐6‐chloro‐2‐methoxyacridine fluorescence quenching as compared to EF 1 or EF 1 (‐δ)+ E. coli δ by energization of the vesicles with NADH, while EF 1 (‐δ)+chloroplast δ was ineffective. All ‘mixed’ combinations were probably reconstitutively active only by plugging the protonic leak through the exposed F o (structural reconstitution) rather than by catalytic activity. Nevertheless, the cross‐reconstitution is stunning in view of the weak sequence similarity between chloroplast δ and E. coli δ. It favors a role of δ as a conformational transducer rather than as a proton conductor between F 0 and F 1 .