Mobilon Cristiane, de Toledo Marcelo Augusto Szymanski, Paganelli Fernanda Laroza, dos Santos Clelton Aparecido, de Pace Fernanda, de Paiva Jacqueline Boldrin, Stehling Eliana Guedes, Nakazato Gerson, Balieiro Aline Gambaro, Airoldi Flavia Pereira da Silva, Reis Francisco de Assis Machado, da Silveira Wanderley Dias
Department of Genetics, Evolution and Bioagents, State University of Campinas, Campinas, SP, Brazil.
Protein Pept Lett. 2013 Feb;20(2):133-9. doi: 10.2174/092986613804725316.
Shigella flexneri is a Gram-negative bacillus that is responsible for a severe form of dysentery called Shigellosis, which mainly affects children and the elderly in both underdeveloped and developed countries. Pathogenic S. flexneri strains possess a large virulence plasmid that codes for effector proteins that are required for the entry and spread of the bacteria into colonocytes. Among these proteins is the translocator IpaC, which plays an important role in the invasion process; IpaC is implicated in pore formation in the host cell membrane and induces cytoskeletal rearrangements in macrophages and epithelial cells, thereby promoting bacterial entry. The ability of IpaC to insert onto the plasma membrane is due to a large nonpolar region of the protein structure. This characteristic also renders difficulties in recovery and purification when the protein is expressed in E. coli. Several works have considered different methodologies for the improved production and purification of IpaC. Herein, we propose an alternative method that is based on changes in the induction temperature and extraction buffer to facilitate the accumulation of high yields of soluble proteins for their further processing and ultimate use in biotechnological approaches.
福氏志贺菌是一种革兰氏阴性杆菌，可引发一种严重的痢疾，称为志贺氏菌病，主要影响欠发达国家和发达国家的儿童及老年人。致病性福氏志贺菌菌株拥有一个大型毒力质粒，该质粒编码效应蛋白，这些蛋白是细菌进入并在结肠细胞中传播所必需的。这些蛋白中包括转运蛋白IpaC，它在侵袭过程中起重要作用；IpaC参与宿主细胞膜上的孔形成，并诱导巨噬细胞和上皮细胞中的细胞骨架重排，从而促进细菌进入。IpaC插入质膜的能力归因于该蛋白质结构中的一个大的非极性区域。当该蛋白在大肠杆菌中表达时，这一特性也给回收和纯化带来困难。几项研究考虑了不同的方法来改进IpaC的生产和纯化。在此，我们提出一种替代方法，该方法基于诱导温度和提取缓冲液的变化，以促进高产率可溶性蛋白的积累，以便进一步加工并最终用于生物技术方法。
