V-ATPase

Vacuolar-type ATPase (V-ATPase) is a highly conserved evolutionarily ancient

The Archaea-type ATPase (A-ATPase) is a related group of ATPases found in archaea that often work as an ATP synthase. It forms a clade V/A-ATPase with V-ATPase. Most members of either group shuttle protons (H⁺
), but a few members have evolved to use sodium ions (Na⁺
) instead.

Roles played by V-ATPases

V-ATPases are found within the membranes of many organelles, such as

V-ATPases also play a significant role in cell morphogenesis development. Disruption of the gene vma-1 gene which encodes for the catalytic subunit (A) of the enzyme severely impairs the rate of growth, differentiation, and the capacity to produce viable spores in fungus Neurospora crassa. [6]

Structure

It has been suggested that this section should be split into articles titled V/A-ATPase subunit C, V/A-ATPase subunits E and G, V/A-ATPase subunit H and V/A-ATPase subunit d/C. (discuss) (September 2020)

The yeast V-ATPase is the best characterized. There are at least thirteen subunits identified to form a functional V-ATPase complex, which consists of two domains. The subunits belong to either the V_o domain (membrane associated subunits, lowercase letters on the figure) or the V₁ domain (peripherally associated subunits, uppercase letters on the figure).

The V₁ includes eight subunits, A-H, with three copies of the catalytic A and B subunits, three copies of the stator subunits E and G, and one copy of the regulatory C and H subunits. In addition, the V₁ domain also contains the subunits D and F, which form a central rotor axle.^[7] The V₁ domain contains tissue-specific subunit isoforms including B, C, E, and G. Mutations to the B1 isoform result in the human disease distal renal tubular acidosis and sensorineural deafness.

The V_o domain contains six different subunits, a, d, c, c', c", and e, with the stoichiometry of the c ring still a matter of debate with a decamer being postulated for the tobacco hornworm (Manduca sexta) V-ATPase. The mammalian V_o domain contains tissue-specific isoforms for subunits a and d, while yeast V-ATPase contains two organelle-specific subunit isoforms of a, Vph1p, and Stv1p. Mutations to the a3 isoform result in the human disease infantile malignant osteopetrosis, and mutations to the a4 isoform result in distal renal tubular acidosis, in some cases with sensorineural deafness.

The V₁ domain is responsible for ATP hydrolysis, whereas the V_o domain is responsible for proton translocation. ATP hydrolysis at the catalytic nucleotide binding sites on subunit A drives rotation of a central stalk composed of subunits D and F, which in turn drives rotation of a barrel of c subunits relative to the a subunit. The complex structure of the V-ATPase has been revealed through the structure of the M. Sexta and Yeast complexes that were solved by single-particle cryo-EM and negative staining, respectively.^[8][9][10] These structures have revealed that the V-ATPase has a 3-stator network, linked by a collar of density formed by the C, H, and a subunits, which, while dividing the V₁ and V_o domains, make no interactions with the central rotor axle formed by the F, D, and d subunits. Rotation of this central rotor axle caused by the hydrolysis of ATP within the catalytic AB domains results in the movement of the barrel of c subunits past the a subunit, which drives proton transport across the membrane. A stoichiometry of two protons translocated for each ATP hydrolyzed has been proposed by Johnson.^[11]

In addition to the structural subunits of yeast V-ATPase, associated proteins that are necessary for assembly have been identified. These associated proteins are essential for V_o domain assembly and are termed Vma12p, Vma21p, and Vma22p.^{[12][13][14][15]} Two of the three proteins, Vma12p and Vma22p, form a complex that binds transiently to Vph1p (subunit a) to aid its assembly and maturation.^{[14][16][17][18]} Vma21p coordinates assembly of the V_o subunits as well as escorting the V_o domain into vesicles for transport to the Golgi.^[19]

V₁

The V₁ domain of the V-ATPase is the site of ATP hydrolysis. Unlike V_o, the V₁ domain is hydrophilic.^[5] This soluble domain consists of a hexamer of alternating A and B subunits, a central rotor D, peripheral stators G and E, and regulatory subunits C and H. Hydrolysis of ATP drives a conformational change in the six A|B interfaces and with it rotation of the central rotor D. Unlike with the ATP synthase, the V₁ domain is not an active ATPase when dissociated.

Subunit C

V-ATPase (Vacuolar-ATPase) C represents the

The C subunit plays an essential role in controlling the assembly of V-ATPase, acting as a flexible stator that holds together the catalytic (V1) and membrane (VO) sectors of the enzyme .^[22] The release of subunit C from the ATPase complex results in the dissociation of the V1 and Vo subcomplexes, which is an important mechanism in controlling V-ATPase activity in cells. Essentially, by creating a high electrochemical gradient and low pH, this powers the enzyme to create more ATP.

Subunits E, G

These related subunits make up the stalk(s) of A/V-ATPase. They are important in assembly, and may function as pushrods in activity. E has a cap to connect to A/B, while G does not.^[20] They likely evolved from a single protein by gene duplication.^[23]

Subunit H

Subunit H, is only involved in activity and not in assembly. This subunit also acts as an inhibitor of free V1 subunits; it stops ATP hydrolysis when V1 and Vo are dissociated.^[24]

V_o

The V_o domain is responsible for proton translocation. Unlike the F-type ATP synthase, the V_o domain generally transports protons against their own concentration gradient. Rotation of the V_o domain transports the protons in movement coordinated with the V₁ domain, which is responsible for ATP hydrolysis. The V_o domain is hydrophobic and composed of several dissociable subunits.^[5] These subunits are present in the V_o domain to make this a functional proton translocase; they are described below.

Subunit a/I

The 116kDa subunit (or subunit a) and subunit I are found in the Vo or Ao complex of V- or A-ATPases, respectively. The 116kDa subunit is a transmembrane glycoprotein required for the assembly and proton transport activity of the ATPase complex. Several isoforms of the 116kDa subunit exist, providing a potential role in the differential targeting and regulation of the V-ATPase for specific organelles.

The function of the 116-kDa subunit is not defined, but its predicted structure consists of 6–8 transmembranous sectors, suggesting that it may function similar to subunit a of FO.

Subunit d/C

Subunit d in V-ATPases, called subunit C in A-ATPases, is a part of the Vo complex. They fit onto the middle of the c ring, so are thought to function as a rotor. There are two versions of this subunit in eukaryotes, d/d1 and d2.^[25]

In mammals, d1 (ATP6V0D1) is the ubiquitously expressed version and d2 (ATP6V0D2) is expressed in specific cell types only.^[25]

Subunit c

Similar to the F-type ATP synthase, the transmembrane region of the V-ATPase includes a ring of membrane-spanning subunits that are primarily responsible for proton translocation. Dissimilar from the F-type ATP synthase, however, the V-ATPase has multiple related subunits in the c-ring; in fungi such as yeast there are three related subunits (of varied stoichiometry) and in most other eukaryotes there are two.

V-ATPase assembly

Yeast V-ATPases fail to assemble when any of the genes that encode subunits are deleted except for subunits H and c".^[26][27][28] Without subunit H, the assembled V-ATPase is not active,^[13][29] and the loss of the c" subunit results in uncoupling of enzymatic activity.^[27]

The precise mechanisms by which V-ATPases assembly are still controversial, with evidence suggesting two different possibilities. Mutational analysis and in vitro assays have shown that preassembled V_o and V₁ domains can combine to form one complex in a process called independent assembly. Support for independent assembly includes the findings that the assembled V_o domain can be found at the vacuole in the absence of the V₁ domain, whereas free V₁ domains can be found in the cytoplasm and not at the vacuole.^[30][31] In contrast, in vivo pulse-chase experiments have revealed early interactions between V_o and V₁ subunits, to be specific, the a and B subunits, suggesting that subunits are added in a step-wise fashion to form a single complex in a concerted assembly process.^[32]

V-ATPase evolution

A relatively new technique called

V-ATPases are known to be specifically inhibited by macrolide antibiotics, such as concanamycin (CCA) and balifomycin A1.[37] In vivo regulation of V-ATPase activity is accomplished by reversible dissociation of the V₁ domain from the V_o domain. After initial assembly, both the insect Manduca sexta and yeast V-ATPases can reversibly disassemble into free V_o and V₁ domains after a 2- to 5-minute deprivation of glucose.^[30] Reversible disassembly may be a general mechanism of regulating V-ATPase activity, since it exists in yeast and insects. Reassembly is proposed to be aided by a complex termed RAVE (regulator of H⁺
-ATPase of vacuolar and endosomal membranes).^[38] Disassembly and reassembly of V-ATPases does not require new protein synthesis but does need an intact microtubular network.^[39]

Human diseases

Osteopetrosis

Osteopetrosis is generic name that represents a group of heritable conditions in which there is a defect in osteoclastic bone resorption. Both dominant and recessive osteopetrosis occur in humans.^[40][41] Autosomal dominant osteopetrosis shows mild symptoms in adults experiencing frequent bone fractures due to brittle bones.^[40] A more severe form of osteopetrosis is termed autosomal recessive infantile malignant osteopetrosis.^[41][42][43] Three genes that are responsible for recessive osteopetrosis in humans have been identified. They are all directly involved in the proton generation and secretion pathways that are essential for bone resorption. One gene is carbonic anhydrase II (CAII), which, when mutated, causes osteopetrosis with renal tubular acidosis(type 3).^[44] Mutations to the chloride channel ClC7 gene also lead to both dominant and recessive osteopetrosis.^[40] Approximately 50% of patients with recessive infantile malignant osteopetrosis have mutations to the a3 subunit isoform of V-ATPase.^{[42] [45][46]} In humans, 26 mutations have been identified in V-ATPase subunit isoform a3, found in osteoclasts, that result in the bone disease autosomal recessive osteopetrosis.^{[42][41][45][47]}

Distal renal tubular acidosis (dRTA)

The importance of V-ATPase activity in renal proton secretion is highlighted by the inherited disease distal

The term V_o has a lowercase letter "o" (not the number "zero") in subscript. The "o" stands for oligomycin, which binds to the homologous region in F-ATPase. It is worth noting that the human gene notations at NCBI designate it as "zero" rather than the letter "o". For example, the gene for the human c subunit of Vo is listed in NCBI gene database as "ATP6V0C" (with a zero), rather than "ATP6VOC" (with an "o"). Many pieces of literature make this mistake as well.

See also

• ATP synthase
• F-ATPase
• Na+/K+-ATPase

References