CRYSTAL-STRUCTURE OF THE EPSILON-SUBUNIT OF THE PROTON-TRANSLOCATING ATP SYNTHASE FROM ESCHERICHIA-COLI

Background: Proton-translocating ATP synthases convert the energy gene rated from photosynthesis or respiration into ATP. These enzymes, term ed F0F1-ATPases, are structurally highly conserved. In Escherichia col i, F0F1-ATPase consists of a membrane portion, F-0, made up of three d ifferent polypeptides (a, b and c) and an F-1 portion comprising five different polypeptides in the stoichiometry alpha(3) beta(3) gamma del ta epsilon. The minor subunits gamma, delta and epsilon are required f or the coupling of proton translocation with ATP synthesis; the epsilo n subunit is in close contact with the alpha, beta, gamma and c subuni ts. The structure of the epsilon subunit provides clues to its essenti al role in this complex enzyme. Results: The structure of the E. coli F0F1-ATPase epsilon subunit has been solved at 2.3 Angstrom resolution by multiple isomorphous replacement. The structure, comprising residu es 2-136 of the polypeptide chain and 14 water molecules, refined to a n R value of 0.214 (R-free = 0.288). The molecule has a novel fold wit h two domains. The N-terminal domain is a beta sandwich with two five- stranded sheets. The C-terminal domain is formed from two alpha helice s arranged in an antiparallel coiled-coil. A series of alanine residue s from each helix form the central contacting residues in the helical domain and can be described as an 'alanine zipper'. There is an extens ive hydrophobic contact region between the two domains providing a sta ble interface. The individual domains of the crystal structure closely resemble the structures determined in solution by NMR spectroscopy. C onclusions: Sequence alignments of a number of epsilon subunits from d iverse sources suggest that the C-terminal domain, which is absent in some species, is not essential for function. In the crystal the N-term inal domains of two epsilon subunits make a close hydrophobic interact ion across a crystallographic twofold axis. This region has previously been proposed as the contact surface between the epsilon and gamma su bunits in the complete F-1-ATPase complex. In the crystal structure, w e observe what is apparently a stable interface between the two domain s of the epsilon subunit, consistent with the fact that the crystal an d solution structures are quite similar despite close crystal packing. This suggests that a gross conformational change in the epsilon subun it, to transmit the effect of proton translocation to the catalytic do main, is unlikely, but cannot be ruled out.