Purified subunit delta of chloroplast coupling factor CF1 reconstitutes photophosphorylation in partially CF1-depleted membranes.
The ATP synthase of chloroplasts consists of the proton channel, CF0, and the catalytic part, CF1, which carries nucleotide-binding sites on subunits alpha and beta. The still poorly understood interaction between CF0 and the catalytic sites on CF1 is mediated by the smaller subunits gamma, delta and epsilon of CF1. We investigated the ability of purified delta to block proton leakage through CF0 channels after their exposure by removal of the CF1 counterpart. Thylakoids were partially depleted of CF1 by EDTA treatment. This increased their proton permeability and thereby reduced the rate of photophosphorylation. Subunit delta was isolated and purified by FPLC [Engelbrecht, S. and Junge, W. (1987) FEBS Lett. 219, 321-325]. Addition of delta to EDTA-treated thylakoids reconstituted high rates of phenazine-methosulfate-mediated photophosphorylation. Since delta does not interact with nucleotides by itself, the reconstitution was due to a reduction of the proton leakage through open CF0 channels. The molar ratio of purified delta over exposed CF0, which started to elicit this effect, was 3:1. However, if delta was added together with purified CF1 lacking delta, in a 1:1 molar ratio, the relative amount over exposed CF0 was as low as 0.06. This corroborated our previous conclusion [Lill, H., Engelbrecht, S., Schönknecht, G. and Junge, W. (1986) Eur. J. Biochem. 160, 627-634] that only a very small fraction of exposed CF0 was actually proton-conducting but with a very high unit conductance. CF1 including delta was apparently rebound preferentially to open CF0 channels. Although the ability of delta to control proton conduction through CF0 was evident, it remains to be established whether delta acts as a gated proton valve or as a conformational transducer in the integral CF0CF1 ATPase.
D000255:Adenosine Triphosphate / Q000302:isolation & purification; D002384:Catalysis; D004492:Edetic Acid; D046911:Macromolecular Substances; D008565:Membrane Proteins / Q000302:isolation & purification*; D010785:Photophosphorylation; D010940:Plant Proteins; D010944:Plants / Q000032:analysis; D006180:Proton-Translocating ATPases / Q000302:isolation & purification* / Q000502:physiology; D011522:Protons
D046911:Macromolecular Substances; D008565:Membrane Proteins; D010940:Plant Proteins; D011522:Protons; C042085:thylakoid membrane phosphoprotein, plant; D000255:Adenosine Triphosphate; D004492:Edetic Acid; D006180:Proton-Translocating ATPases