Structure of the mitochondrial F1 ATPase at 9-A resolution.

Abstract

The soluble portion (F1 ATPase) of the mitochondrial ATP-synthesizing system is a multisubunit enzyme of molecular weight 380,000. It is composed of five different subunits, a, .3, y, 8, and e. The subunit stoichiometry is not known but there are strong suggestions that it is a333y8e. We have determined the three-dimensional structure of the F1 ATPase of rat liver mitochondria to 9-A resolution by using x-ray diffraction techniques. The molecule appears to be formed by two equivalent halves, each formed by three regions ofapproximately equal size. These regions form a distorted hexagonal or octahedral arrangement. None of the regions form closed symmetrical trimers in the complex. It is proposed that, if the subunit stoichiometry is a3p3y8e, the major subunits exist in at least two different environments in the complex. In this arrangement, the different copies of the major subunits are functionally not equivalent. This observation appears to offer a natural explanation of the complicated binding and labeling data of F1 ATPases.The main pathway for the production ofATP in living organisms involves the phosphorylation of ADP by inorganic phosphate. It is clear from thermodynamic considerations that the levels of cellular ATP needed to perform biochemical phosphorylations, adenylations, and active transport should be above those determined by the hydrolytic equilibrium ATP + H20 = ADP + Pi. The ATP-synthesizing systems utilize the energy provided by the oxidation of substrates or by the capture of light quanta for the production ofsuch ATP levels. Coupling between the energy-yielding processes and the phosphorylation reaction is considered to be effected by an electrochemical proton gradient established across a biological membrane separating two compartments (1). The ATP-synthesizing system is then, according to this view, a proton-translocating ATPase.ATP-synthesizing complexes of similar structure are found in mitochondria, chloroplasts, and bacteria (for review, see refs. 2-6). They are large membrane-bound proteins composed of three morphologically distinct parts: a membrane sector, a connecting stalk, and a head piece or F1 ATPase.In this paper, we report an x-ray crystallographic study ofthe F1 ATPase from rat liver mitochondria (7). This protein has a molecular weight of380,000 and is composed offive nonidentical subunits a, A3, y, 8, and e (8) whose molecular weights estimated from NaDodSO4 gels are a, 62,500; (3, 57,000; 'y, 36,000; 8, 12,000; and E, 7,500. The stain intensity ofthese subunits in NaDodSO4 gels is compatible with a subunit stoichiometry of a3f33y5E (8). However, the subunit stoichiometry of F1 ATPases is still an unsolved problem and stoichiometries of the type a2322y282E2 are also considered possible in this system (for review, see ref. 6). The work described here provides information about the quaternary structure of the F1 ATPase of rat liver and in addition addresses the problem of the subunit stoichiometry of the enzyme. Our structure determination was carried out using the ...

Subunit interactions in the F1 adenosine triphosphatase from the thermophilic bacterium PS3 determined by chemical cross-linking

“…In one possible model the three a subunits and the three P subunits could be placed at opposite ends of the hexagon, as we have proposed for the F1 ATPase of E. coli (21). This asymmetric model is consistent with the results of X-ray diffraction (28,29) and reconstitution studies (30) of the F1 ATPase of rat liver mitochondria. However, it should be noted that Liinsdorf et al (31) using monoclonal antibodies to the a and P subunits of the F1 ATPase of E. coli have found by electron microscopy that a maximum of three molecules of immunoglobulin were bound to the enzyme, and in a symmetrical manner.…”

Cooperative binding of manganese(II) to chloroplast coupling factor 1 detected by NMR proton relaxation enhancement

“…This conclusion is supported by the observation that rapid turnover of the enzyme requires the binding of substrate to two of the three active sites (Grubmeyer & Penefsky, 1981; Cross et al, 1982). X-ray diffraction analysis does not support a simple 3-fold symmetry of subunits, suggesting structural inequivalence of the /3 subunits (Amzel et al, 1982).…”

The proton-ATPase of bacteria and mitochondria

“…Nevertheless an ~3fl37 complex will have inherent asymmetry in its ~-7 or fl-~/interfaces. Amzel et al (1982) have recently determined the three-dimensional structure of mitochondrial F 1 in the presence of ATP to 9 ~ resolution by X-ray diffraction techniques. For an e3]~37~e stoichiometry, they conclude an inherent asymmetry of ~ and ]?…”