Photosynthetic ATPases: purification, properties, subunit isolation and function

Abstract

Photosynthetic coupling factor ATPases (F1-ATPases) generally censist of five subunits named α, β, γ, δ and ε in order of decreasing apparent molecular weight. The isolated enzyme has a molecular weight of between 390,000 to 400,000, with the five subunits probably occurring in a 3:3:1:1:1 ratio. Some photosynthetic F1 ATPases are inactive as isolated and require treatment with protease, heat or detergent in order to elicit ATPase activity. This activity is sensitive to inhibition by free divalent cations and appears to be more specific for Ca(2+) vs. Mg(2+) as the metal ion substrate chelate. This preference for Ca(2+) can be explained by the higher inhibition constant for inhibition of ATPase activity by free Ca(2+). Methods for the assay of a Mg-dependent ATPase activity have recently been described. These depend on the presence of organic solvents or detergents in the reaction mixture for assay. The molecular mechanism behind the expression of either the Ca- or Mg-ATPase activities is unknown. F1-ATPases function to couple proton efflux from thylakoid membranes or chromatophores to ATP synthesis. The isolated enzyme may thus also be assayed for the reconstitution of 'coupling activity' to membranes depleted of coupling factor 1.The functions of the five subunits in the complex have been deduced from the results of chemical modification and reconstitution studies. The δ subunit is required for the functional binding of the F1 to the F0. The active site is probably contained in the β (and α) subunit(s). The proposed functions for the γ and ε subunits are, however, still matters of controversy. Coupling factors from a wide variety of species including bacteria, algae, C3 and C4 plants, appear to be immunologically related. The β subunits are the most strongly related, although the α and γ subunits also show significant immunological cross-reactivity. DNA sequence analyses of the genes for the β subunit of CF1 have indicated that the primary sequence of this polypeptide is highly conserved. The genes for the polypeptides of CF1 appear to be located in two cellular compartments. The α, β and ε subunits are coded for on chloroplast DNA, whereas the γ and δ subunits are probably nuclear encoded. Experiments involving protein synthesis by isolated chloroplasts or protein synthesis in the presence of inhibitors specific for one or the other set of ribosomes in the cell suggest the existence of pools of unassembled CF1 subunits. These pools, if they do exist in vivo, probably make up no greater than 1% of the total CF1 content of the cell.