Determining the Stoichiometry of Photosynthetic Phosphorylation

Abstract

AFTER the discovery of non-cyclic photosynthetic phosphorylation¹, investigations in other laboratories^2,3 confirmed the observation that one ATP molecule was formed for each pair of electrons (2e) transferred from water to TPN⁺ or ferricyanide. Later work with these electron acceptors^4,5 and with quinones⁶ added support to the belief that the limit of the P : 2e ratio (μmoles ATP formed per two electrons transferred) is unity. Tris-HCl was used in this early work, and more recent studies using this buffer have confirmed the reported pH optimum of 7.8–8.0 and the P : 2e ratio of 1.0 (ref. 7). With the introduction of new buffers⁸ and the use of pH values between 8.6 and 8.9, overall P : 2e ratios greater than 1.0 have been observed^7–9. Lynn and Brown¹⁰, using a different incubation procedure, claim to have obtained high P : 2e ratios with the tris buffer, and Shavit and Avron¹¹ have obtained ratios greater than 1.0 with tris-HCl. On the other hand, Ramirez et al.¹² have never-observed P : 2e ratios greater than 1.0 although the pH optima for ferricyanide and TPN⁺ reduction were 7.7 and 8.5, respectively, in the presence of tris, but were both 8.5 in the presence of ‘Tricine’ (N-tris (hydroxymethyl) methylglycine). The H⁺ : e (ratio of protons taken up to electrons transported; the value of this H⁺ : e ratio should equal the number of sites of energy conservation (ATP formation) in the electron transport path) varies strongly with pH showing a maximum value of 6 at pH 6.0 using diquat (N, N-ethylene-2,2-dipyridilium dibromide) as the electron acceptor; a similar variation with pH occurred in ferricyanide reduction with a maximum H⁺ : e = 4 (ref. 13). An H⁺ : e ratio of 5 was observed with chloranil as the electron acceptor⁹. Many different P : 2e and H⁺ : e ratios have thus been observed depending on the pH, the electron acceptor and the buffer.