Estimation of the H+/H- ratio of the reaction catalysed by the nicotinamide nucleotide transhydrogenase in chromatophores from over-expressing strains of Rhodospirillum rubrum and in liposomes inlaid with the purified bovine enzyme

Two strains of Rhodospirillum rubrum were constructed in which, by a gene dosage effect, the transhydrogenase activity of isolated chromatophores was increased 7-10-fold and 15-20-fold, respectively. The H+/H- ratio (the ratio of protons translocated per hydride ion equivalent transferred from NADPH to an NAD+ analogue, acetyl pyridine adenine dinucleotide), determined by a spectroscopic technique, was approximately 1.0 for chromatophores from the over-expressing strains, but was only approximately 0.6 for wild-type chromatophores. Highly-coupled proteoliposomes were prepared containing purified transhydrogenase from beef-heart mitochondria. Using the same technique, the H+/H- ratio was close to 1.0 for these proteoliposomes. It is suggested that the mechanistic H+/H- ratio is indeed unity, but that a low ratio is obtained in wild-type chromatophores because of inhomogeneity in the vesicle population.