Lemieux M Joanne, Huang Yafei, Wang Da Neng
Skirball Institute for Biomolecular Medicine, New York University School of Medicine, 540 First Ave., 3-5, New York, NY 10016, USA.
J Electron Microsc (Tokyo). 2005;54 Suppl 1:i43-6. doi: 10.1093/jmicro/54.suppl_1.i43.
The major facilitator superfamily represents the largest group of secondary active membrane transporters in prokaryotic and eukaryotic cells. They transport a vast variety of substrates, presumably via similar mechanisms, yet the details of these mechanisms remain unclear. Here we report the 3.3 A resolution structure of a member of this superfamily--GlpT, the glycerol-3-phosphate transporter from the E. coli inner membrane, in the absence of a substrate. The antiporter mediates the exchange of glycerol-3-phosphate for inorganic phosphate across the membrane. Its N- and C-terminal domains exhibit a pseudo 2-fold symmetry along an axis perpendicular to the membrane. Eight of the twelve transmembrane alpha-helices are arranged around a centrally located substrate translocation pore that is closed off at the periplasmic surface. Present at the beginning of the pore are two arginine residues that presumably comprise the substrate-binding site which is accessible only from the cytosol, suggesting an inward-facing conformation for the transporter. The central loop connecting the N- and C-terminal domains is partially disordered and exhibits reduced susceptibility to trypsin in the presence of substrate, indicating conformational changes. We propose that GlpT operates via a single binding-site, alternating-access mechanism.
主要易化子超家族是原核细胞和真核细胞中最大的一类次级主动膜转运蛋白。它们通过相似的机制转运各种各样的底物，然而这些机制的细节仍不清楚。在此，我们报道了该超家族一个成员——GlpT（大肠杆菌内膜的甘油-3-磷酸转运蛋白）在无底物情况下分辨率为3.3埃的结构。该反向转运蛋白介导甘油-3-磷酸与无机磷酸跨膜交换。其N端和C端结构域沿垂直于膜的轴呈现假二重对称。十二个跨膜α螺旋中的八个围绕位于中心的底物转运孔排列，该孔在周质表面封闭。孔的起始处有两个精氨酸残基，推测它们构成仅从胞质溶胶可及的底物结合位点，这表明转运蛋白处于向内的构象。连接N端和C端结构域的中央环部分无序，并且在有底物存在时对胰蛋白酶的敏感性降低，表明存在构象变化。我们提出GlpT通过单一结合位点交替访问机制发挥作用。
