Uhlin U, Cox G B, Guss J M
Department of Biochemistry, University of Sydney, NSW, Australia.
Structure. 1997 Sep 15;5(9):1219-30. doi: 10.1016/s0969-2126(97)00272-4.
Proton-translocating ATP synthases convert the energy generated from photosynthesis or respiration into ATP. These enzymes, termed F0F1-ATPases, are structurally highly conserved. In Escherichia coli, F0F1-ATPase consists of a membrane portion, F0, made up of three different polypeptides (a, b and c) and an F1 portion comprising five different polypeptides in the stoichiometry alpha 3 beta 3 gamma delta epsilon. The minor subunits gamma, delta and epsilon are required for the coupling of proton translocation with ATP synthesis; the epsilon subunit is in close contact with the alpha, beta, gamma and c subunits. The structure of the epsilon subunit provides clues to its essential role in this complex enzyme.
The structure of the E. coli F0F1-ATPase epsilon subunit has been solved at 2.3 A resolution by multiple isomorphous replacement. The structure, comprising residues 2-136 of the polypeptide chain and 14 water molecules, refined to an R value of 0.214 (Rfree = 0.288). The molecule has a novel fold with two domains. The N-terminal domain is a beta sandwich with two five-stranded sheets. The C-terminal domain is formed from two alpha helices arranged in an antiparallel coiled-coil. A series of alanine residues from each helix form the central contacting residues in the helical domain and can be described as an 'alanine zipper'. There is an extensive hydrophobic contact region between the two domains providing a stable interface. The individual domains of the crystal structure closely resemble the structures determined in solution by NMR spectroscopy.
Sequence alignments of a number of epsilon subunits from diverse sources suggest that the C-terminal domain, which is absent in some species, is not essential for function. In the crystal the N-terminal domains of two epsilon subunits make a close hydrophobic interaction across a crystallographic twofold axis. This region has previously been proposed as the contact surface between the epsilon and gamma subunits in the complete F1-ATPase complex. In the crystal structure we observe what is apparently a stable interface between the two domains of the epsilon subunit, consistent with the fact that the crystal and solution structures are quite similar despite close crystal packing. This suggests that a gross conformational change in the epsilon subunit, to transmit the effect of proton translocation to the catalytic domain, is unlikely, but cannot be ruled out.
质子转运ATP合酶将光合作用或呼吸作用产生的能量转化为ATP。这些酶被称为F0F1 - ATP酶，在结构上高度保守。在大肠杆菌中，F0F1 - ATP酶由一个膜部分F0和一个F1部分组成，F0由三种不同的多肽（a、b和c）组成，F1部分按化学计量比α3β3γδε包含五种不同的多肽。γ、δ和ε亚基是质子转运与ATP合成偶联所必需的；ε亚基与α、β、γ和c亚基紧密接触。ε亚基的结构为其在这种复合酶中的重要作用提供了线索。
通过多同晶置换法以2.3埃的分辨率解析了大肠杆菌F0F1 - ATP酶ε亚基的结构。该结构由多肽链的2 - 136位残基和14个水分子组成，精修后的R值为0.214（Rfree = 0.288）。该分子具有一个新颖的折叠结构，包含两个结构域。N端结构域是一个由两个五链片层组成的β三明治结构。C端结构域由两个反平行卷曲螺旋排列的α螺旋形成。每个螺旋上的一系列丙氨酸残基形成螺旋结构域中的中心接触残基，可被描述为一个“丙氨酸拉链”。两个结构域之间有一个广泛的疏水接触区域，提供了一个稳定的界面。晶体结构的各个结构域与通过核磁共振光谱在溶液中测定的结构非常相似。
来自不同来源的多个ε亚基的序列比对表明，一些物种中不存在的C端结构域对功能并非必不可少。在晶体中，两个ε亚基的N端结构域在一个晶体学二重轴上形成紧密的疏水相互作用。该区域先前被认为是完整F1 - ATP酶复合物中ε亚基和γ亚基之间的接触表面。在晶体结构中，我们观察到ε亚基的两个结构域之间显然存在一个稳定的界面，这与尽管晶体堆积紧密但晶体结构和溶液结构非常相似的事实一致。这表明ε亚基发生重大构象变化以将质子转运的效应传递到催化结构域的可能性不大，但不能排除这种可能性。
