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...

Archaeal A(1)A(O) ATP synthase/ATPase operons are highly conserved among species and comprise at least nine genes encoding structural proteins. However, all A(1)A(O) ATPase preparations reported to date contained only three to six subunits and, therefore, the study of this unique class of secondary energy converters is still in its infancy. To improve the quality of A(1)A(O) ATPase preparations, we chose the hyperthermophilic, methanogenic archaeon Methanococcus jannaschii as a model organism. Individual subunits of the A(1)A(O) ATPase from M. jannaschii were produced in E. coli, purified, and antibodies were raised. The antibodies enabled the development of a protocol ensuring purification of the entire nine-subunit A(1)A(O) ATPase. The ATPase was solubilized from membranes of M. jannaschii by Triton X-100 and purified to apparent homogeneity by sucrose density gradient centrifugation, ion exchange chromatography, and gel filtration. Electron micrographs revealed the A(1) and A(O) domains and the central stalk, but also additional masses which could represent a second stalk. Inhibitor studies were used to demonstrate that the A(1) and A(O) domains are functionally coupled. This is the first description of an A(1)A(O) ATPase preparation in which the two domains (A(1) and A(O)) are fully conserved and functionally coupled.

ATPases with unusual membrane-embedded rotor subunits were found in both F(1)F(0) and A(1)A(0) ATP synthases. The rotor subunit c of A(1)A(0) ATPases is, in most cases, similar to subunit c from F(0). Surprisingly, multiplied c subunits with four, six, or even 26 transmembrane spans have been found in some archaea and these multiplication events were sometimes accompanied by loss of the ion-translocating group. Nevertheless, these enzymes are still active as ATP synthases. A duplicated c subunit with only one ion-translocating group was found along with "normal" F(0) c subunits in the Na(+) F(1)F(0) ATP synthase of the bacterium Acetobacterium woodii. These extraordinary features and exceptional structural and functional variability in the rotor of ATP synthases may have arisen as an adaptation to different cellular needs and the extreme physicochemical conditions in the early history of life.

Archaea are a heterogenous group of microorganisms that often thrive under harsh environmental conditions such as high temperatures, extreme pHs and high salinity. As other living cells, they use chemiosmotic mechanisms along with substrate level phosphorylation to conserve energy in form of ATP. Because some archaea are rooted close to the origin in the tree of life, these unusual mechanisms are considered to have developed very early in the history of life and, therefore, may represent first energy-conserving mechanisms. A key component in cellular bioenergetics is the ATP synthase. The enzyme from archaea represents a new class of ATPases, the A₁A₀ ATP synthases. They are composed of two domains that function as a pair of rotary motors connected by a central and peripheral stalk(s). The structure of the chemically-driven motor (A₁) was solved by small-angle X-ray scattering in solution, and the structure of the first A₁A₀ ATP synthases was obtained recently by single particle analyses. These studies revealed novel structural features such as a second peripheral stalk and a collar-like structure. In addition, the membrane-embedded electrically-driven motor (A₀) is very different in archaea with sometimes novel, exceptional subunit composition and coupling stoichiometries that may reflect the differences in energy-conserving mechanisms as well as adaptation to temperatures at or above 100°C.