Structure and Subunit Arrangement of the A-type ATP Synthase Complex from the Archaeon Methanococcus jannaschii Visualized by Electron Microscopy

Abstract

In Archaea, bacteria, and eukarya, ATP provides metabolic energy for energy-dependent processes. It is synthesized by enzymes known as A-type or F-type ATP synthase, which are the smallest rotatory engines in nature (Yoshida, M., Muneyuki, E., and Hisabori, T. ATP synthases/ATPases are present in every life form and are the most important enzymes for the energy metabolism of the cell (1). They catalyze the formation of ATP at the expense of the transmembrane electrochemical ion gradient. They arose from a common ancestor that underwent structural and functional changes leading to three distinct classes of A 1 A 0 , 1 F 1 F 0 , and V 1 V 0 ATP synthases/ATPases. The V-type ATPases, found in organelles of eukaryotes, lost their ability to synthesize ATP. Their function is to create steep ion gradients at the expense of ATP hydrolysis (2). Archaea contain ATPases, the A 1 A 0 ATP synthases, that are structurally similar to V 1 V 0 ATPases but synthesize ATP like the F-type ATPases. The genomic sequences available today show that the overall subunit composition of the A 1 A 0 ATP synthases is very similar to the V 1 V 0 ATPases. For example, the A 1 A 0 ATP synthases contain duplicated and even triplicated K subunits (proteolipids) (3, 4). The A 1 A 0 ATP synthase has at least nine subunits (A 3 B 3 CDEFHIK X ), but the actual subunit stoichiometry, especially regarding the proteolipid subunits K in A ATPases, is different in various organisms (12, 6, 4, or as suggested by genomic data, only 1 (5)). The A 1 A 0 ATP synthase is composed of a water-soluble A 1 ATPase and an integral membrane subcomplex, A 0 . ATP is synthesized or hydrolyzed on the A 1 headpiece consisting of an A 3 B 3 domain, and the energy that is provided for or released during that process is transmitted to the membrane-bound A 0 domain (4). The energy coupling between the two active domains occurs via the so-called stalk part, an assembly proposed to be composed of the subunits C, D, and F (6, 7). Insight in the molecular structure of the A-type ATP synthases comes from small-angle x-ray scattering data of the A 1 ATPase from Methanosarcina mazei Gö1 whose A 1 domain is made up of the five different subunits, A 3 B 3 CDF (8). The data have shown that the hydrated A 1 ATPase is rather elongated with a headpiece of 10 ϫ 9.4 nm in dimension and a stalk of ϳ8.4 nm in length. A comparison of the central stalk of this A 1 complex with bacterial F 1 and eukaryotic V 1 ATPase indicates different lengths of the stalk domain (8, 9). Image processing of electron micrographs of negatively stained A 1 ATPase from M. mazei Gö1 (7) has revealed that the headpiece consists of a pseudohexagonal arrangement of six masses surrounding a seventh mass. These barrel-shaped masses of ϳ3.2 and 2.8 nm in diameter and 7.5 and 5.0 nm in length, which consist of the major subunits A and B, are arranged in an alternating manner (7). The hexagonal barrel of subunits A and B encloses a cavity of ϳ2.3 nm in the middle in which part of the central stalk is asymmetrically located...

The Stator Complex of the A1A0-ATP Synthase—Structural Characterization of the E and H Subunits

“…11 The biophysical characterization of the EH complex provided here is consistent with this structural requirement. First, the complex is highly α helical, including a region of coiled coil (see above).…”

“…For reasons of simplicity, the coiled-coil nature of the interaction between subunit H and the N-terminal domain of subunit E (E N ) has been omitted from the model. The collar observed in EM images of the intact A-ATPase 11 is formed in our model by the hydrophilic domain of subunit I plus contributions from the N-terminal domains of the EH heterodimer that interact with the I subunit hydrophilic domain. For details, see the text.…”

“…In the bacterial F-ATPase, a single peripheral stalk is found, which is composed of the membrane-anchored b subunits together with the δ subunit bound on the top of the F 1 . 8 The situation is less clear in the A-and V-ATPases; electron microscopy studies have suggested the presence of multiple peripheral stators in the related enzymes [9][10][11] and studies with individual polypeptides have led to the proposal that dimeric species of subunits E 12 and H 13 (or G 14 ) are structural elements of the A-(or V-) ATPase peripheral stalks. A role of the cytoplasmic domain of A-ATPase subunit I 15 (or V-ATPase subunit a 16 ) as peripheral stator has also been proposed.…”

Domain Architecture of the Stator Complex of the A1A0-ATP Synthase from Thermoplasma acidophilum

“…whereas two heterodimeric EH complexes are thought to form the peripheral stalk attachment to A 0 seen in electron microscopy reconstructions (5,6). In the A 0 domain (subunits CIK x ), the K subunits (proteolipids) form a ring that is linked to the central stalk by the C subunit, whereas the cytoplasmic N-terminal domain of the I subunit probably mediates the binding of the EH peripheral stalks to A 0 , as suggested for the bacterial A/V-type enzyme (7).…”

Stoichiometry and Localization of the Stator Subunits E and Gin Thermus thermophilus H+-ATPase/Synthase

“…Part of the stator is the peripheral stalk, which counteracts the rotation of the catalytic A 3 B 3 head with respect to the spinning central rotor in all types of ATPases. It has been visualized by electron microscopy (EM) in various F-ATPases (9, 12-14), in V-ATPases (8, 10), and in other eubacterial/archaeal V/AATPases (7,15,16). These studies suggest that F-ATPases contain a single peripheral stalk, A-ATPases contain two, and eukaryotic V-ATPases might contain up to three.…”