Molecular Physiology and Biological Physics, University of Virginia School of Medicine, Charlottesville, Virginia 22908, USA.
Department of Molecular and Cellular Biology, University of Guelph, Guelph, Ontario N1G 2W1, Canada.
Nature. 2018 Jan 18;553(7688):361-365. doi: 10.1038/nature25190. Epub 2018 Jan 10.
O-antigens are cell surface polysaccharides of many Gram-negative pathogens that aid in escaping innate immune responses. A widespread O-antigen biosynthesis mechanism involves the synthesis of the lipid-anchored polymer on the cytosolic face of the inner membrane, followed by transport to the periplasmic side where it is ligated to the lipid A core to complete a lipopolysaccharide molecule. In this pathway, transport to the periplasm is mediated by an ATP-binding cassette (ABC) transporter, called Wzm-Wzt. Here we present the crystal structure of the Wzm-Wzt homologue from Aquifex aeolicus in an open conformation. The transporter forms a transmembrane channel that is sufficiently wide to accommodate a linear polysaccharide. Its nucleotide-binding domain and a periplasmic extension form 'gate helices' at the cytosolic and periplasmic membrane interfaces that probably serve as substrate entry and exit points. Site-directed mutagenesis of the gates impairs in vivo O-antigen secretion in the Escherichia coli prototype. Combined with a closed structure of the isolated nucleotide-binding domains, our structural and functional analyses suggest a processive O-antigen translocation mechanism, which stands in contrast to the classical alternating access mechanism of ABC transporters.
O-抗原是许多革兰氏阴性病原体的细胞表面多糖，有助于逃避先天免疫反应。广泛存在的 O-抗原生物合成机制涉及在内膜细胞质面合成脂锚定聚合物，然后转运到周质侧，在那里与脂 A 核心连接完成脂多糖分子。在该途径中，向周质的转运由称为 Wzm-Wzt 的 ATP 结合盒 (ABC) 转运体介导。在这里，我们展示了来自水生栖热菌的 Wzm-Wzt 同源物在开放构象下的晶体结构。该转运体形成一个足够宽的跨膜通道，可容纳线性多糖。其核苷酸结合域和周质延伸在细胞质和周质膜界面处形成“门螺旋”，可能作为底物进入和退出点。门的定点突变会损害大肠杆菌原型物中 O-抗原的体内分泌。结合分离的核苷酸结合结构域的封闭结构，我们的结构和功能分析表明了一种连续的 O-抗原转运机制，这与 ABC 转运体的经典交替访问机制形成对比。
