Alerasol Masoome, Mousavi Gargari Seyed Latif, Nazarian Shahram, Bagheri Samane
Dept. of Biology, Shahed University, Tehran, Iran.
Depent of Biology, Imam Hussein University, Tehran, Iran.
Iran Biomed J. 2014;18(4):212-218. doi: 10.6091/ibj.1344.2014.
Enterotoxigenic Escherichia coli (ETEC) strains are the major causes of diarrheal disease in humans and animals. Colonization factors and enterotoxins are the major virulence factors in ETEC pathogenesis. For the broad-spectrum protection against ETEC, one could focus on colonization factors and non-toxic heat labile as a vaccine candidate.
A fusion protein is composed of a major fimbrial subunit of coli surface antigen 3, and the heat-labile B subunit (LTB) was constructed as a chimeric immunogen. For optimum level expression of protein, the gene was synthesized with codon bias of E. coli. Also, recombinant protein was expressed in E. coli BL21DE3. ELISA and Western tests were carried out for determination of antigen and specificity of antibody raised against recombinant protein in animals. The anti-toxicity and anti-adherence properties of the immune sera against ETEC were also evaluated.
Immunological analyses showed the production of high titer of specific antibody in immunized mice. The built-in LTB retains native toxin properties which were approved by GM1 binding assay. Pre-treatment of the ETEC cells with anti-sera significantly decreased their adhesion to Caco-2 cells.
The results indicated the efficacy of the recombinant chimeric protein as an effective immunogen inducing strong humoral response. The designated chimer would be an interesting prototype for a vaccine and worthy of further investigation.
产肠毒素大肠杆菌（ETEC）菌株是人和动物腹泻病的主要病因。定植因子和肠毒素是ETEC发病机制中的主要毒力因子。为了对ETEC提供广谱保护，可以将重点放在定植因子和无毒热不稳定蛋白作为疫苗候选物上。
构建一种融合蛋白，其由大肠杆菌表面抗原3的主要菌毛亚基和热不稳定B亚基（LTB）组成，作为嵌合免疫原。为了实现蛋白的最佳表达水平，根据大肠杆菌的密码子偏好性合成基因。此外，重组蛋白在大肠杆菌BL21DE3中表达。进行ELISA和Western检测，以确定动物体内针对重组蛋白产生的抗体的抗原性和特异性。还评估了免疫血清对ETEC的抗毒性和抗黏附特性。
免疫分析显示免疫小鼠产生了高滴度的特异性抗体。内置的LTB保留了天然毒素特性，这通过GM1结合试验得到证实。用抗血清预处理ETEC细胞可显著降低其对Caco-2细胞的黏附。
结果表明重组嵌合蛋白作为一种能诱导强烈体液反应的有效免疫原具有有效性。指定的嵌合体将是一种有趣的疫苗原型，值得进一步研究。
