Physiology of V-ATPases.

Protons migrate much faster than other ions through water, ice and water-lined membrane channels because they participate in hydrogen bonding and H/H20 exchange. Similarly, hydrogen bonding enables protons with amino, carbonyl, phosphoryl and sulfonyl residues to influence critically the charge, conformation and stability of proteins. Therefore, it is not surprising that regulation of proton concentration, or pH, is an essential requirement in biological systems. It is no surprise either that enzymes which regulate proton concentration (i.e. proton pumps) should have evolved or that evolution should have used these enzymes further, for energization of biological membranes. At present there appear to be three classes of ATP-hydrolyzing proton pumps, or HATPases, which were dubbed P-ATPases, F-ATPases and V-ATPases, by Pederson and Carafoli (1987). H-translocating P-ATPases, as well as the Na/K-ATPase of plasma membranes and the Ca-ATPase of sarcoplasmic reticulum, form phosphoaspartyl intermediates and are inhibited by the phosphate analogue orthovanadate. F-ATPases are the proton-translocating ATP synthases of mitochondria, chloroplasts and bacterial plasma membranes and are inhibited by azide. V-ATPases (V-type ATPases or vacuolar H-ATPases) are the proton-translocating ATPases best known as acidifiers of cytoplasmic vesicles and vacuoles. During reaction, they separate protons from gegenions, thus producing an electrical potential difference, A^P, across the membrane in which they reside. A ^ in turn can drive other ion movements, leading directly to acidification or alkalization and secondarily to salt transport and water movement. V-ATPases are inhibited by bafilomycin (in nanomolar concentrations) and A'-ethylmaleimide (in micromolar concentrations). In this paper the transport work that is accomplished when specific channels and porters are present in V-ATPase-energized membranes will be discussed using an intuitive thermodynamic analysis. Martin (1992) discusses similar cases using ionic circuit analysis. The two approaches provide complementary frameworks for understanding how the (output) compartment receiving protons from a V-ATPase can be rendered acidic, neutral or even alkaline, depending upon the selectivity of other carriers (porters) and channels present, upon the strength of the gegenions and upon the cellular and compartmental geometry. Encouragingly, both types of analysis lead to similar conclusions. Biochemical studies on V-ATPases were initiated on vacuolar membranes of fungi and plants and on clathrin-coated vesicles and other components of endomembrane systems (reviewed by Sze, 1985; Mellman etal. 1986; Nelson, 1988, 1989; Forgac, 1989; Stone et