Living Systems Institute, University of Exeter, Exeter, UK.
Faculty of Health and Life Sciences, University of Exeter, Exeter, UK.
Methods Mol Biol. 2024;2778:291-310. doi: 10.1007/978-1-0716-3734-0_18.
Secretin proteins form pores in the outer membranes of Gram-negative bacteria, and as such provide a means of transporting a wide variety of molecules out of or in to the cell. They are important components of several different bacterial secretion systems, surface filament assembly machineries, and virus assembly complexes. Despite accommodating a diverse assortment of molecules, including virulence factors, folded proteins, and whole viruses, the secretin family of proteins is highly conserved, particularly in their membrane-embedded β-barrel domain. We describe here a protocol for the expression, purification and cryo-EM structural determination of the pIV secretin from the Ff family of filamentous bacteriophages.
分泌蛋白在革兰氏阴性菌的外膜上形成孔道，因此提供了一种将各种分子从细胞内或细胞外运输的方法。它们是几种不同的细菌分泌系统、表面丝组装机器和病毒组装复合物的重要组成部分。尽管分泌蛋白家族可以容纳包括毒力因子、折叠蛋白和整个病毒在内的各种分子，但它们在其膜嵌入式β桶结构域中高度保守。我们在这里描述了从丝状噬菌体 Ff 家族的 pIV 分泌蛋白的表达、纯化和冷冻电镜结构测定的方案。
