Vacuolar ATPases function

In the cell, the vacuolar ATPase functions exclusively as an ATP hydrolysis-driven proton pump. This functional preference of the vacuolar ATPase seems not to be due to a fundamental structural difference compared with the ATP synthase because it has been shown that the V-ATPase can be reversed to synthesize ATP in vitro, albeit with very low efficiency (Hirata et al., 2000). 

One of the first inteins discovered was found in the 119-kDa precursor to a subunit of a vacuolar ATPase of yeast.a,c In this 50-kDa intein Thr 72, His 75, and His 197 may have catalytic functions.d The intein is spliced out to form the 69-kDa subunit. 

The F-, V-, and A-ATPases constitute a family of ATP hydrolysis-driven ion pumps which are found in Archaea, eubacteria, simple eukaryotes such as yeast, and higher eukaryotes including plants and mammals. The family of ion pumps is divided into three subfamilies the F-ATPases (which function mainly as ATP synthases), the vacuolar ATPases (which function solely as ATP hydrolysis-driven ion pumps) and the Archaeal A-type ATPases (whose function can be either in the direction of ATP synthesis or hydrolysis). All three members of the family are evolutionarily related, and it is believed that the three subfamilies have arisen from a common ancestor. 

Proton-ATPases can be divided into three classes a) The plasma-membrane type, which operates via a phosphoenzyme intermediate and therefore is part of the P-ATPase superfamily. These proton pumps evolved from a common ancestor of the Ca" and Na pumps and are structurally distinct from the other two families of proton pumps (1-3). b) The eubacterial-type F-ATPases that are present in eubacteria, mitochondria and chloroplasts (3,4). c) The vacuolar-type V-ATPases that are present in archaebacteria and the vacuolar system of eukaryotic cells (2-6). F and V-ATPases are structurally and functionally related and have evolved from a common ancestral enzyme (3,4). This relationship was established from a wealth of sequence information regarding F-ATPases and by more recent studies on V-ATPases. The divergent pathways by which F and V-ATPases have evolved were recently elucidated by pai lel studies in several laboratories (3). It is the piupose of this communication to discuss aspects pertinent to the evolution of CFq-CFi, which is the F-ATPase functioning in photosynthesis. 

Arata, Y., Nish, T., Kawasaki-Nishi, S., Shao, E., Wilkens, S. and Forgac, M. Structure, subunit function and regulation of the coated vesicle and yeast vacuolar H - ATPases. Biochim. Biophys. Acta 1555 71-74, 2002. 

Mattsson JP, Li X, Peng S-B, Nilsson F, Adersen P, Lundberg LG, Stone DK, Keeling DJ. 2000. Properties of three isoforms of the 116-kDa Subunit of vacuolar H+-ATPase from a single vertebrate species. Cloning, gene expression and protein characterization of functionally distinct isoforms in Gallus domesticus. Euro J Bioch 267 4115-26. 

An H+ electrochemical gradient (ApH+) provides the energy required for active transport of all classical neurotransmitters into synaptic vesicles. The Mg2+-dependent vacuolar-type H+-ATPase (V-ATPase) that produces this gradient resides on internal membranes of the secretory pathway, in particular endosomes and lysosomes (vacuole in yeast) as well as secretory vesicles (Figure 3). In terms of both structure and function, this pump resembles the F-type ATPases of bacteria, mitochondria and chloroplasts, and differs from the P-type ATPases expressed at the plasma membrane of mammalian cells (e.g., the Na+/K+-, gastric H+/K+-and muscle Ca2+-ATPases) (Forgac, 1989 Nelson, 1992). The vacuolar and F0F1... 

The ease of hydrolysis can vary enormously in some cases hydrolysis is so easy that inadvertent loss of the protecting group during chromatography becomes a nuisance (such as acetals of a, f3-unsaturated ketones). On the other hand acetals may be so robust that forcing conditions (mineral acid and heat) are required. For example, substrates that contain a basic amino function — even if it is remote — can be quite resistant to hydrolysis, because protonation first takes place at the more basic amino function,18 The resultant positive charge repels the second Q-protonation required to set in motion the hydrolysis. An acid-catalysed hydrolysis of a basic acetal that required refluxing with 6 M HC1 in acetone for 6-10 h19 is illustrated in Scheme 2,3, Perhaps the mildest conditions employ pyridinium p-toluenesulfonate (PPTS, pKa 5.2) in methanol or aqueous acetone exemplified by the transacetalisation reaction taken from a synthesis of the vacuolar H+-ATPase inhibitor Bafilomycin Ai [Scheme 2.4, 20... 

Eide, D. J., Bridgham, J. T, Zhao, Z., and Mattoon, J. R. (1993). The vacuolar H -ATPase of Saccharomyces cerevisiae is required for efficient copper detoxification, mitochondrial function, and iron metabolism. Mol. Gen. Genet. 241, 447 56. 

However, proteins in mammalian cells manage to escape from quality control in the ER to adopt alternative or dual topology in different intracellular membrane compartments. There are an increasing number of examples of proteins that are expressed in different topological forms with different functions. For example, ductin was found in two different orientations in cellular membranes, one of which serves as the subunit of the vacuolar H -ATPase and the other serves as a component of the microsomal connexin channel of gap... 

Stevens TH, Forgac M. Structure, function and regulation of the vacuolar (H )-ATPase. Annu Rev Cell Dev Biol 1997 13 779-808. 

Garrett-Engele R, Moilanen B., and Cyert M.S. 1995. Calcineurin, the Ca /calmodulin-dependent protein phosphatase is essential in yeast mutants with cell integrity defects and in mutants that lack a functional vacuolar H -ATPase. Mol Cell Biol 15 4103-4114. 

Vacuoles also have a second principal function they stock metabolites before their use. In fact, tliey contain a quarter of the pool of the amino acids of the cell, including a lot of arginine as well as 5-adenosyl methionine. In this organelle, there is also potassium, adenine, isoguanine, uric acid and polyphosphate crystals. These are involved in the fixation of basic amino acids. Specific permeases ensure the transport of these metabolites across the vacuolar membrane. An ATPase linked to the tonoplast furnishes the necessary energy for the movement of stocked compounds against the concentration gradient. It is different from the plasmic membrane ATPase, but also produces a proton efflux. 

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