Structure and Properties of the Vacuolar (H+)-ATPases (1999)

The vacuolar (H+)-ATPases (or V-ATPases) are a family of ATP-dependent proton pumps that are responsible for acidification of intracellular compartments in eukaryotic cells (1Stevens T.H. Forgac M. Annu. Rev. Cell Dev. Biol. 1997; 13: 779-808Crossref PubMed Scopus (520) Google Scholar, 2Bowman B.J. Bowman E.J. Brambl Marzluf The Mycota III: Biochemistry and Molecular Biology. Springer-Verlag, Berlin1996: 57-83Google Scholar, 3Forgac M. FEBS Lett. 1998; 440: 258-263Crossref PubMed Scopus (118) Google Scholar). Acidification of vacuolar compartments plays an important role in a variety of cellular processes, particularly membrane traffic processes. In receptor-mediated endocytosis, acidification of endosomes serves as a signal that activates release of internalized ligands (such as low density lipoprotein and insulin) from their receptors. This uncoupling allows unoccupied receptors to recycle to the cell surface where they can be reutilized, whereas the ligands are targeted to lysosomes for degradation (1Stevens T.H. Forgac M. Annu. Rev. Cell Dev. Biol. 1997; 13: 779-808Crossref PubMed Scopus (520) Google Scholar). Acidification of endosomes is also required for the formation of endosomal carrier vesicles (which are involved in moving ligands along the endocytic pathway (4Clague M.J. Urbe S. Aniento F. Gruenberg J. J. Biol. Chem. 1994; 269: 21-24Abstract Full Text PDF PubMed Google Scholar)) and in activating fusion between the endosomal membrane and internalized envelope viruses (such as influenza virus) that is necessary for viral infection (5White J.M. Science. 1992; 258: 917-924Crossref PubMed Scopus (649) Google Scholar). In intracellular targeting of newly synthesized lysosomal enzymes, vacuolar acidification plays a very similar role to that observed for endocytosis. Thus, lysosomal enzymes bound to the mannose 6-phosphate receptor are delivered to a late recycling compartment where the low pH causes dissociation, allowing receptors to recycle to the trans-Golgi and the enzymes to be targeted to lysosomes (6Sahagian G.G. Novikoff P.M. Arias I. Boyer J. Fausto N. Jakoby W. Schachter D. Shafritz D. The Liver: Biology and Pathobiology. 3rd Ed. Raven Press, New York1994: 275-291Google Scholar). Disruption of vacuolar acidification in yeast also causes a perturbation in vacuolar targeting (7Klionsky D.J. Nelson H. Nelson N. J. Biol. Chem. 1992; 267: 3416-3422Abstract Full Text PDF PubMed Google Scholar), although the molecular basis of this effect is not yet clear. In lysosomes and other digestive organelles, like the central vacuole of yeast, the low pH is required to support the activity of acid hydrolases involved in degradation of macromolecules. In both central vacuoles and secretory vesicles, such as synaptic vesicles and chromaffin granules, the proton gradient or membrane potential generated by the V-ATPase is utilized to drive the coupled transport of small molecules into the vesicle lumen (8Moriyama Y. Madea M. Futai M. J. Exp. Biol. 1992; 172: 171-178Crossref PubMed Google Scholar, 9Anraku Y. Umemoto N. Hirata R. Ohya Y. J. Bioenerg. Biomembr. 1992; 24: 395-406Crossref PubMed Scopus (64) Google Scholar). In addition to their role in intracellular compartments, V-ATPases have also been shown to play an important role in the plasma membrane of various specialized cells. Thus V-ATPases in the apical membrane of renal intercalated cells function in renal acidification (10Gluck S.L. J. Bioenerg. Biomembr. 1992; 24: 351-360Crossref PubMed Scopus (53) Google Scholar), whereas plasma membrane V-ATPases in macrophages and neutrophils assist in pH homeostasis (11Swallow C.J. Grinstein S. Sudsbury R.A. Rotstein O.D. J. Cell. Physiol. 1993; 157: 453-460Crossref PubMed Scopus (60) Google Scholar). Osteoclasts (12Chatterjee D. Chakraborty M. Leit M. Neff L. Jamsa-Kellokumpu S. Fuchs R. Baron R. Proc. Natl. Acad. Sci. U. S. A. 1992; 89: 6257-6261Crossref PubMed Scopus (133) Google Scholar) and tumor cells (13Martinez-Zaguilan R. Lynch R. Martinez G. Gillies R. Am. J. Physiol. 1993; 265: C1015-C1029Crossref PubMed Google Scholar) are able to target V-ATPases to the plasma membrane where they create an acidic extracellular environment that is necessary for bone resorption or tumor metastasis, respectively. V-ATPases in the midgut of insects have been shown to establish a membrane potential across the apical membrane that drives K+ transport via an electrogenic H+/K+ antiporter (14Wieczorek H. Putzenlechner M. Zeiske W. Klein U. J. Biol. Chem. 1991; 266: 15340-15347Abstract Full Text PDF PubMed Google Scholar). Finally, using the specific V-ATPase inhibitor bafilomycin (15Bowman E.J. Siebers A. Altendorf K. Proc. Natl. Acad. Sci. U. S. A. 1988; 85: 7972-7976Crossref PubMed Scopus (1589) Google Scholar), V-ATPases have been implicated in apoptosis (16Gottlieb R.A. Giesing H.A. Zhu J.Y. Engler R.L. Babior B.M. Proc. Natl. Acad. Sci. U. S. A. 1995; 92: 5965-5968Crossref PubMed Scopus (231) Google Scholar). It is thus clear that V-ATPases serve a wide variety of roles in eukaryotic cells. Our current model for the structure of the V-ATPases is shown in Fig. 1. The subunit composition of the V-ATPase, together with the yeast genes encoding the V-ATPase subunits, is shown in Table I. The V-ATPases are composed of two functional domains (1Stevens T.H. Forgac M. Annu. Rev. Cell Dev. Biol. 1997; 13: 779-808Crossref PubMed Scopus (520) Google Scholar, 2Bowman B.J. Bowman E.J. Brambl Marzluf The Mycota III: Biochemistry and Molecular Biology. Springer-Verlag, Berlin1996: 57-83Google Scholar, 3Forgac M. FEBS Lett. 1998; 440: 258-263Crossref PubMed Scopus (118) Google Scholar). The V1 domain is a 570-kDa peripheral complex responsible for hydrolysis of ATP. V1 is composed of eight different subunits of molecular mass 70–14 kDa (subunits A–H), with three copies each of the A and B subunits and single copies of the remaining subunits (17Arai H. Terres G. Pink S. Forgac M. J. Biol. Chem. 1988; 263: 8796-8802Abstract Full Text PDF PubMed Google Scholar). The A and B subunits both participate in nucleotide binding, with the catalytic site located on the A subunit (1Stevens T.H. Forgac M. Annu. Rev. Cell Dev. Biol. 1997; 13: 779-808Crossref PubMed Scopus (520) Google Scholar, 2Bowman B.J. Bowman E.J. Brambl Marzluf The Mycota III: Biochemistry and Molecular Biology. Springer-Verlag, Berlin1996: 57-83Google Scholar, 3Forgac M. FEBS Lett. 1998; 440: 258-263Crossref PubMed Scopus (118) Google Scholar). The V0 domain is a 260-kDa integral complex that is responsible for proton translocation across the membrane. V0 contains five different subunits of molecular mass 100–17 kDa (subunits a, d, c, c′, and c“), with six copies of the c/c′ subunits and single copies of the other subunits (17Arai H. Terres G. Pink S. Forgac M. J. Biol. Chem. 1988; 263: 8796-8802Abstract Full Text PDF PubMed Google Scholar).Table ISubunit composition of V-ATPasesDomainSubunitBovineMrYeast MrYeast geneaIn yeast, disruption of vma genes encoding the V-ATPase subunits leads to loss of V-ATPase activity and a conditional lethal phenotype in which cells are able to grow at acidic pH but not at neutral pH or in the presence of elevated extracellular Ca2+ (1, 9).Subunit functionV1A7369VMA1Catalytic site, regulation (?)B5857VMA2Noncatalytic site, targeting (?)C4042VMA5Activity, assemblyD3432VMA8Activity, assemblyE3327VMA4Activity, assemblyF1414VMA7Activity, assemblyG1513VMA10Activity, assemblyH5054VMA13Activity (not assembly)V0a10095VPH1/STV1H+transport, assembly, targetingd3836VMA6Activity, assemblyc1717VMA3H+ translocation, DCCD sitec′1717VMA11H+translocation, DCCD site (?)c“1923VMA16H+translocationa In yeast, disruption of vma genes encoding the V-ATPase subunits leads to loss of V-ATPase activity and a conditional lethal phenotype in which cells are able to grow at acidic pH but not at neutral pH or in the presence of elevated extracellular Ca2+ (1Stevens T.H. Forgac M. Annu. Rev. Cell Dev. Biol. 1997; 13: 779-808Crossref PubMed Scopus (520) Google Scholar, 9Anraku Y. Umemoto N. Hirata R. Ohya Y. J. Bioenerg. Biomembr. 1992; 24: 395-406Crossref PubMed Scopus (64) Google Scholar). Open table in a new tab The V-ATPases are structurally and evolutionarily related to the F-ATPases (or ATP synthases) of mitochondria, chloroplasts, and bacteria (18Weber J. Senior A. Biochim. Biophys. Acta. 1997; 1319: 19-58Crossref PubMed Scopus (395) Google Scholar, 19Fillingame R.H. J. Exp. Biol. 1997; 200: 217-224Crossref PubMed Google Scholar, 20Cross R.L. Duncan T.M. J. Bioenerg. Biomembr. 1996; 28: 403-408Crossref PubMed Scopus (67) Google Scholar) that normally function to couple the synthesis of ATP to the movement of protons down their electrochemical gradient. This relationship is evident both in their overall structure (3Forgac M. FEBS Lett. 1998; 440: 258-263Crossref PubMed Scopus (118) Google Scholar) and in the sequence of the nucleotide binding and proteolipid subunits (subunit c) (2Bowman B.J. Bowman E.J. Brambl Marzluf The Mycota III: Biochemistry and Molecular Biology. Springer-Verlag, Berlin1996: 57-83Google Scholar). Despite this similarity, however, the V- and F-ATPases differ in many important respects, particularly in their function and regulation. An important goal is to determine the structural basis for these differences between the V- and F-ATPases Comparison of electron microscopic images of the V-ATPase fromNeurospora and the F-ATPase of mitochondria revealed that although both contained a globular head attached to the membrane by a central stalk, the V-ATPase had additional projections emanating from the base of the stalk (21Dschida W.J. Bowman B.J. J. Biol. Chem. 1992; 267: 18783-18789Abstract Full Text PDF PubMed Google Scholar). More recent images of the V-ATPase fromClostridium fervidus have revealed the presence of a second or peripheral stalk (22Boekema E.J. Ubbink-Kok T. Lolkema J.S. Brisson A. Konings W.N. Proc. Natl. Acad. Sci. U. S. A. 1997; 94: 14291-14293Crossref PubMed Scopus (91) Google Scholar), similar to that reported for the F-ATPase ofEscherichia coli (23Wilkens S. Capaldi R.A. Nature. 1998; 393: 29Crossref PubMed Scopus (135) Google Scholar). Negatively stained images of the bovine coated vesicle V-ATPase reveal an even more extensive secondary stalk. 1S. Wilkens and M. Forgac, unpublished observations. This peripheral stalk is crucial for the current rotary model of ATP-driven proton transport by both the F- and V-ATPases (20Cross R.L. Duncan T.M. J. Bioenerg. Biomembr. 1996; 28: 403-408Crossref PubMed Scopus (67) Google Scholar, 24Junge W. Sabbert D. Engelbrecht S. Ber. Bunsen-ges. Phys. Chem. 1996; 100: 2014-2019Crossref Google Scholar, 54Vik S.B. Antonio B.J. J. Biol. Chem. 1994; 269: 30364-30369Abstract Full Text PDF PubMed Google Scholar), discussed in detail below. The nucleotide binding sites of the V-ATPase are located on two subunits: the 70-kDa A and 60-kDa B subunits. Several lines of evidence suggest that the catalytic nucleotide binding sites are located on the A subunit. First, ATP-protectable modification of the A subunit by such reagents as N-ethylmaleimide, 7-chloro-4-nitrobenz-2-oxa-1,3-diazole (NBD-Cl), and 2-azido-ATP correlates well with loss of activity (3Forgac M. FEBS Lett. 1998; 440: 258-263Crossref PubMed Scopus (118) Google Scholar, 25Zhang J. Vasilyeva E. Feng Y. Forgac M. J. Biol. Chem. 1995; 270: 15494-15500Abstract Full Text Full Text PDF PubMed Scopus (39) Google Scholar). Second, the A subunit contains all of the consensus nucleotide binding sequences found in the catalytic nucleotide binding subunit (β) of the F-ATPase (2Bowman B.J. Bowman E.J. Brambl Marzluf The Mycota III: Biochemistry and Molecular Biology. Springer-Verlag, Berlin1996: 57-83Google Scholar). For the β subunit, the location of these sequences has been identified from the x-ray crystal structure of F1 (26Abrahams J.P. Leslie A.G. Lutter R. Walker J.E. Nature. 1994; 370: 621-628Crossref PubMed Scopus (2743) Google Scholar, 27Bianchet M.A. Hullihen J. Pedersen P.L. Amzel L.M. Proc. Natl. Acad. Sci. U. S. A. 1998; 95: 11065-11070Crossref PubMed Scopus (224) Google Scholar). Although no high resolution structural data are yet available for the V-ATPases, mutation of conserved residues in these sequences in the A subunit leads to significant losses of activity (28Liu J. Kane P.M. Biochemistry. 1996; 35: 10938-10948Crossref PubMed Scopus (55) Google Scholar, 29Liu Q. Leng X.H. Newman P. Vasilyeva E. Kane P.M. Forgac M. J. Biol. Chem. 1997; 272: 11750-11756Abstract Full Text Full Text PDF PubMed Scopus (76) Google Scholar, 30MacLeod K.J. Vasilyeva E. Baleja J.D. Forgac M. J. Biol. Chem. 1998; Full Text Full Text PDF PubMed Scopus Google Scholar), as observed for the β subunit (18Weber J. Senior A. Biochim. Biophys. Acta. 1997; 1319: 19-58Crossref PubMed Scopus (395) Google Scholar). that the V- and the F-ATPases is the of the V-ATPases to such (1Stevens T.H. Forgac M. Annu. Rev. Cell Dev. Biol. 1997; 13: 779-808Crossref PubMed Scopus (520) Google Scholar, 2Bowman B.J. Bowman E.J. Brambl Marzluf The Mycota III: Biochemistry and Molecular Biology. Springer-Verlag, Berlin1996: 57-83Google Scholar, 3Forgac M. FEBS Lett. 1998; 440: 258-263Crossref PubMed Scopus (118) Google Scholar). of the bovine V-ATPase A subunit has been identified as the responsible for the of the V-ATPases to reagents Y. Forgac M. J. Biol. Chem. 1992; 267: Full Text PDF PubMed Google Scholar). is located in the consensus sequence from the crystal structure of the of ATP bound at the catalytic site (26Abrahams J.P. Leslie A.G. Lutter R. Walker J.E. Nature. 1994; 370: 621-628Crossref PubMed Scopus (2743) Google Scholar, 27Bianchet M.A. Hullihen J. Pedersen P.L. Amzel L.M. Proc. Natl. Acad. Sci. U. S. A. 1998; 95: 11065-11070Crossref PubMed Scopus (224) Google Scholar). has also been shown to a with a second in the A subunit, to of V-ATPase activity Y. Forgac M. J. Biol. Chem. 1994; 269: Full Text PDF PubMed Google Scholar). is located in the domain of the A subunit, and formation is to activity by the of domains with nucleotide as in the crystal structure of F1 (26Abrahams J.P. Leslie A.G. Lutter R. Walker J.E. Nature. 1994; 370: 621-628Crossref PubMed Scopus (2743) Google Scholar). The role of formation in regulation of V-ATPase activity is discussed below. with the nucleotide binding sites on the A subunit, the nucleotide binding sites on the B subunit to be The evidence for this is as First, the B subunit not the consensus sequence shown to be for ATP hydrolysis by the F- and V-ATPases (2Bowman B.J. Bowman E.J. Brambl Marzluf The Mycota III: Biochemistry and Molecular Biology. Springer-Verlag, Berlin1996: 57-83Google Scholar). Second, mutation of residues to be at the nucleotide binding sites on the B subunit to in activity Q. Kane P.M. Newman Forgac M. J. Biol. Chem. 1996; Full Text Full Text PDF PubMed Scopus (67) Google Scholar), in to at the catalytic site (28Liu J. Kane P.M. Biochemistry. 1996; 35: 10938-10948Crossref PubMed Scopus (55) Google Scholar, 29Liu Q. Leng X.H. Newman P. Vasilyeva E. Kane P.M. Forgac M. J. Biol. Chem. 1997; 272: 11750-11756Abstract Full Text Full Text PDF PubMed Scopus (76) Google Scholar, 30MacLeod K.J. Vasilyeva E. Baleja J.D. Forgac M. J. Biol. Chem. 1998; Full Text Full Text PDF PubMed Scopus Google Scholar). modification of the site on the B subunit by to loss of activity E. Forgac M. J. Biol. Chem. 1996; Full Text Full Text PDF PubMed Scopus Google Scholar). The function of the sites on the V-ATPases is but recent is of the A subunit (which is to be by the A subunit to the is to or a addition of is observed K.J. Vasilyeva E. Baleja J.D. Forgac M. J. Biol. Chem. 1998; Full Text Full Text PDF PubMed Scopus Google Scholar), that the sites to be to have been the role of other V1 subunits (subunits in activity or of the V-ATPase First, of the genes encoding these subunits in yeast has been shown to to a loss of V-ATPase activity (1Stevens T.H. Forgac M. Annu. Rev. Cell Dev. Biol. 1997; 13: 779-808Crossref PubMed Scopus (520) Google Scholar, 9Anraku Y. Umemoto N. Hirata R. Ohya Y. J. Bioenerg. Biomembr. 1992; 24: 395-406Crossref PubMed Scopus (64) Google Scholar), the V1 domain is no able to the integral V0 domain P.M. I. T. J. Biol. Chem. 1992; 267: Full Text PDF PubMed Google Scholar). to this is subunit which is required for but not assembly, of the V-ATPase complex Hirata R. Umemoto N. Ohya Y. A. T.H. Y. J. Biol. Chem. 1993; Full Text PDF PubMed Google Scholar). In in have a role for many of the V1 subunits in or more by the V1 domain P. J.P. L. D.J. S.B. J. Biol. Chem. 1997; 272: Full Text Full Text PDF PubMed Scopus Google Scholar). Although the V1 domain not K. M. R. Forgac M. J. Biol. Chem. 1992; 267: Full Text PDF PubMed Google Scholar), subunit to this on V1 P. J.P. L. D.J. S.B. J. Biol. Chem. 1997; 272: Full Text Full Text PDF PubMed Scopus Google Scholar). between subunits in the V-ATPase have been First, a between subunits and has been by K. M. R. Forgac M. J. Biol. Chem. 1992; 267: Full Text PDF PubMed Google Scholar). In yeast, the of V1 subunits leads to the formation of a variety of V1 subunits and subunits T.H. D.J. J. Biol. Chem. 1997; 272: Full Text Full Text PDF PubMed Scopus Google Scholar), that these subunits also of the bovine V-ATPase revealed between subunits and and a to be subunit of I. K. N. H. Forgac M. J. Biol. Chem. 265: Full Text PDF PubMed Google Scholar). subunits and are very similar in molecular to subunit c, however, these have and An important is the subunit composition of the central and peripheral observed in the electron microscopic images of the V-ATPase In the the central stalk is composed of the and subunits, whereas the peripheral stalk is of the subunit and the domain of subunit I. R. Capaldi R.A. J. Biol. Chem. 1997; 272: Full Text Full Text PDF PubMed Scopus Google Scholar). The high of subunits and that of these subunits function as the V-ATPase of the subunit, thus of the central stalk E.J. A. Bowman B.J. Biochim. Biophys. Acta. 1995; PubMed Scopus Google Scholar, H. S. Nelson N. Proc. Natl. Acad. Sci. U. S. A. 1995; 92: PubMed Scopus Google Scholar). The subunit composition of the peripheral stalk for the V-ATPase is but subunits and of V1 and subunits and the domain of subunit a in V0 are all potential The V0 domain contains five different of subunits, three of which c′, and are The proteolipid subunits are both to each other R. A. T.H. Y. J. Biol. Chem. 1997; 272: Full Text Full Text PDF PubMed Scopus Google Scholar) and to the subunit of the F-ATPases R.H. J. Exp. Biol. 1997; 200: 217-224Crossref PubMed Google Scholar). The F-ATPase subunit is composed of two a single in that is for proton transport R.H. J. Exp. Biol. 1997; 200: 217-224Crossref PubMed Google Scholar). The V-ATPase subunits and are composed of a single in R. A. T.H. Y. J. Biol. Chem. 1997; 272: Full Text Full Text PDF PubMed Scopus Google Scholar). and of the V-ATPases by and fusion from the F-ATPase subunit N. J. Bioenerg. Biomembr. 1992; 24: PubMed Scopus Google Scholar). are copies of subunit in R.H. J. Exp. Biol. 1997; 200: 217-224Crossref PubMed Google Scholar) and copies of subunits c/c′ in V0 (17Arai H. Terres G. Pink S. Forgac M. J. Biol. Chem. 1988; 263: 8796-8802Abstract Full Text PDF PubMed Google Scholar), these subunits the of to their The of by the subunits in however, is that in and this has been to for the in observed for the V- and F-ATPases R.L. L. FEBS Lett. PubMed Scopus Google Scholar). For both V0 and of a single subunit with DCCD is to proton transport (3Forgac M. FEBS Lett. 1998; 440: 258-263Crossref PubMed Scopus (118) Google Scholar, 19Fillingame R.H. J. Exp. Biol. 1997; 200: 217-224Crossref PubMed Google Scholar), that all the be functional for proton transport to with subunits and c′, subunit is a that contains five with the in the R. A. T.H. Y. J. Biol. Chem. 1997; 272: Full Text Full Text PDF PubMed Scopus Google Scholar). the V-ATPases three different proteolipid subunits to for the is but that be at of each proteolipid subunit in V-ATPase complex R. A. T.H. Y. J. Biol. Chem. 1997; 272: Full Text Full Text PDF PubMed Scopus Google Scholar). of which has the structure that the subunits a with the a and subunits to and the domain of subunit from the membrane R. M. G. F. Altendorf K. J. 1995; PubMed Scopus Google Scholar). Although have not been on the V-ATPase subunit has been observed to that are to A. T. T. T. D. J. J. 1993; PubMed Scopus Google Scholar). In the V-ATPase subunit has been shown to with the of bovine D.J. T. P. K. R. Nature. 1991; PubMed Scopus Google Scholar). suggest the that subunit of the V-ATPases serve roles in the The subunit (subunit of the V-ATPase is a an domain and a domain J. Biol. Chem. 1991; 266: Full Text PDF PubMed Google Scholar). In yeast, the subunit is by two and D. R.A. A. M. D. J. D. J. Biol. Chem. 1992; 267: Full Text PDF PubMed Google Scholar, D. M.A. J. Biol. Chem. 1994; 269: Full Text PDF PubMed Google Scholar). to the central vacuole whereas to other intracellular or endosomes D. M.A. J. Biol. Chem. 1994; 269: Full Text PDF PubMed Google Scholar), that the subunit target the V-ATPase to different compartments in the Although no sequence between suggest like the a subunit of the F-ATPases J. Biol. Chem. 1988; 263: Full Text PDF PubMed Google Scholar, 54Vik S.B. Antonio B.J. J. Biol. Chem. 1994; 269: 30364-30369Abstract Full Text PDF PubMed Google Scholar), the V-ATPase a subunit contains residues that participate in proton translocation X.H. M. Q. Forgac M. J. Biol. Chem. 1996; Full Text Full Text PDF PubMed Scopus Google Scholar, X.H. M. Forgac M. J. Biol. Chem. 1998; Full Text Full Text PDF PubMed Scopus Google Scholar). Although not required for proton transport for the V- X.H. M. Forgac M. J. Biol. Chem. 1998; Full Text Full Text PDF PubMed Scopus Google Scholar) or F-ATPases R.H. J. Biol. Chem. 1997; 272: Full Text Full Text PDF PubMed Scopus Google Scholar), these residues to be in a to proton as in the pH of activity X.H. M. Forgac M. J. Biol. Chem. 1998; Full Text Full Text PDF PubMed Scopus Google Scholar). Thus the a subunit that protons to to the of subunit and to this site to the as has been for the F-ATPase a subunit S.B. Antonio B.J. J. Biol. Chem. 1994; 269: 30364-30369Abstract Full Text PDF PubMed Google Scholar). has also been that the subunit the binding site for the specific V-ATPase inhibitor bafilomycin J. Feng Y. Forgac M. J. Biol. Chem. 1994; 269: Full Text PDF PubMed Google Scholar). In addition to the proteolipid subunits and subunit a, the V0 domain also contains a single of subunit d, a with no N. J. Bioenerg. Biomembr. 1992; 24: PubMed Scopus Google Scholar), which bound to V0 of V1 I. K. N. H. Forgac M. J. Biol. Chem. 265: Full Text PDF PubMed Google Scholar). function has yet been to subunit Although the by which the V-ATPases ATP-dependent proton translocation has not been is to be similar to the rotary for the F-ATPases (20Cross R.L. Duncan T.M. J. Bioenerg. Biomembr. 1996; 28: 403-408Crossref PubMed Scopus (67) Google Scholar, 24Junge W. Sabbert D. Engelbrecht S. Ber. Bunsen-ges. Phys. Chem. 1996; 100: 2014-2019Crossref Google Scholar, 54Vik S.B. Antonio B.J. J. Biol. Chem. 1994; 269: 30364-30369Abstract Full Text PDF PubMed Google Scholar). In the rotary hydrolysis of ATP of the central subunit the of F1 has been by (20Cross R.L. Duncan T.M. J. Bioenerg. Biomembr. 1996; 28: 403-408Crossref PubMed Scopus (67) Google W. Sabbert D. Engelbrecht S. Ber. Bunsen-ges. Phys. Chem. 1996; 100: 2014-2019Crossref Google Scholar), by H. R. M. K. Nature. 1997; PubMed Scopus Google Scholar). The subunit is to be attached to the of subunits in I. R. Capaldi R.A. J. Biol. Chem. 1997; 272: Full Text Full Text PDF PubMed Scopus Google Scholar) that of of the of subunits to subunit a is to the by a peripheral stalk and two to each that protons to and the on subunit S.B. Antonio B.J. J. Biol. Chem. 1994; 269: 30364-30369Abstract Full Text PDF PubMed Google Scholar). the of subunits protons are by the subunit from the and are to the of the membrane the second This the rotary of F1 into a gradient of protons across the membrane. of the yeast V-ATPase that although the V0 domain is able to and target to the central vacuole in the of the V1 subunits, the of of the V0 subunits both of V1 and and targeting of V0 P.M. I. T. J. Biol. Chem. 1992; 267: Full Text PDF PubMed Google Scholar). This that the targeting for the V-ATPase complex in yeast in the V0 which be to the vacuolar membrane. For the of of the V1 subunits of V1 and of V1 to although various V1 are observed these T.H. D.J. J. Biol. Chem. 1997; 272: Full Text Full Text PDF PubMed Scopus Google Scholar). the in this is subunit which is required for activity but not of the V-ATPase complex Hirata R. Umemoto N. Ohya Y. A. T.H. Y. J. Biol. Chem. 1993; Full Text PDF PubMed Google Scholar). of the V-ATPase has also been in where that of V1 and V0 can in the (3Forgac M. FEBS Lett. 1998; 440: 258-263Crossref PubMed Scopus (118) Google Scholar). In yeast, a of and to function in of the V-ATPase but not of the complex (1Stevens T.H. Forgac M. Annu. Rev. Cell Dev. Biol. 1997; 13: 779-808Crossref PubMed Scopus (520) Google Scholar). three of these in the where they to be involved in the of of V0 K.J. T.H. J. Cell Biol. 1998; PubMed Scopus Google Scholar). are in that they to function in of the V-ATPase not in the of other membrane Several subunits have been to necessary for intracellular targeting of the the two of the subunit and are targeted to different intracellular compartments in yeast D. M.A. J. Biol. Chem. 1994; 269: Full Text PDF PubMed Google Scholar). In two of subunit B have been identified (10Gluck S.L. J. Bioenerg. Biomembr. 1992; 24: 351-360Crossref PubMed Scopus (53) Google Scholar). The of B is in all cell whereas the is at high in renal intercalated which are of targeting V-ATPases to the plasma membrane (10Gluck S.L. J. Bioenerg. Biomembr. 1992; 24: 351-360Crossref PubMed Scopus (53) Google Scholar). the B subunit contains targeting necessary to the V-ATPase to the plasma membrane or the two of B the targeted subunits to be It is clear from a variety of that cells are able to different intracellular compartments at different pH and that this is for the by the V-ATPases (1Stevens T.H. Forgac M. Annu. Rev. Cell Dev. Biol. 1997; 13: 779-808Crossref PubMed Scopus (520) Google Scholar). Thus, for endosomes more acidic receptor-mediated to and receptor recycling from the A central in this is cells vacuolar and evidence has been in support of The to be discussed and of the V-ATPase In yeast, has been shown to a of the V-ATPase complex into V1 and V0 domains P.M. J. Biol. Chem. 1995; 270: Full Text Full Text PDF PubMed Google Scholar). This is and of new that the domains are of V1 and is also observed in insects J.P. Klein U. D. H. J. Biol. Chem. 1995; 270: Full Text Full Text PDF PubMed Scopus Google Scholar), and V1 and V0 domains have been identified in cells (3Forgac M. FEBS Lett. 1998; 440: 258-263Crossref PubMed Scopus (118) Google Scholar), that a similar be in these The the of the V-ATPase to be A second to V-ATPase activity in formation between residues at the catalytic site of the an can be between and of the bovine A subunit Y. Forgac M. J. Biol. Chem. 1994; 269: Full Text PDF PubMed Google Scholar), and a significant of the V-ATPase in vesicles in this Y. Forgac M. J. Biol. Chem. 1992; 267: Full Text PDF PubMed Google Scholar). with this mutation of the to in yeast leads to an that is to Q. Leng X.H. Newman P. Vasilyeva E. Kane P.M. Forgac M. J. Biol. Chem. 1997; 272: 11750-11756Abstract Full Text Full Text PDF PubMed Scopus (76) Google Scholar). of the V-ATPase from has also been observed W.J. Bowman B.J. J. Biol. Chem. 1995; 270: Full Text Full Text PDF PubMed Scopus Google Scholar) and has been to be responsible for the of the V-ATPases to The of is but the bovine V-ATPase has been shown to be to the M. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). a mutation in the pathway in yeast leads to vacuolar and this can be by the mutation Kane P.M. J. Biol. Chem. 1997; 272: Full Text Full Text PDF PubMed Scopus Google Scholar), that formation can of V-ATPase activity in that has been to vacuolar acidification is a in the of the V-ATPase N. J. Bioenerg. Biomembr. 1992; 24: PubMed Scopus Google Scholar). A variety of causes a in the of of proton transport and ATP high of ATP and (3Forgac M. FEBS Lett. 1998; 440: 258-263Crossref PubMed Scopus (118) Google Scholar), that the V-ATPase is to in a coupled In has been that an in is to the of the V-ATPase to a vacuolar pH of U. D. L. J. Biol. Chem. 1997; 272: Full Text Full Text PDF PubMed Scopus (55) Google Scholar). with the discussed the cellular are A of low molecular and inhibitor that V-ATPase activity have been identified (10Gluck S.L. J. Bioenerg. Biomembr. 1992; 24: 351-360Crossref PubMed Scopus (53) Google Scholar), that these to vacuolar acidification in In in density have been shown to proton transport across the plasma membrane of renal intercalated cells (10Gluck S.L. J. Bioenerg. Biomembr. 1992; 24: 351-360Crossref PubMed Scopus (53) Google Scholar), although the role of in density in vacuolar pH has not been Finally, V-ATPases are a proton transport for significant acidification to in the important involved in of the membrane potential generated proton transport to be This is by a family of intracellular activity is by (3Forgac M. FEBS Lett. 1998; 440: 258-263Crossref PubMed Scopus (118) Google Scholar). Thus regulation of activity an additional of vacuolar pH in cells. The V-ATPases are a family of ATP-dependent proton pumps that serve a variety of important in the particularly in various membrane traffic are composed of a domain responsible for ATP hydrolysis and an integral domain that proton transport and are structurally related to the ATP of their of that a of are in their activity in