用胶体金标记法观察P195蛋白与人红细胞特异性结合区域  

Observation on specific binding site of P195 protein to human erythrocytes by colloidal gold labeling

作　　者：方军[1,2] 何斌[1,2] 管惟滨[1,2] 

机构地区：[1]第二军医大学基础医学部寄生虫学教研室 [2]第二军医大学军医系

出　　处：《第二军医大学学报》1997年第2期109-112,共4页Academic Journal of Second Military Medical University

基　　金：国家自然科学基金;总后卫生部高技术八五基金

摘　　要：目的：寻找恶性疟原虫裂殖子表面主要蛋白Ｐ１９５中的红细胞结合位点，为设计疫苗阻断裂殖子入侵人红细胞提供实验依据。方法：在大肠杆菌中分８段表达Ｐ１９５蛋白，用镍亲和层析柱分离这些蛋白。各段蛋白经复性后用胶体金标记。标记后的蛋白与人红细胞共孵育，经固定、切片后在电镜下观察。同时将各段蛋白加入到体外培养的人恶性疟原虫的培养基上清中，观察各段蛋白对裂殖子入侵红细胞的影响。结果：Ｐ１９５蛋白中的一段，即Ｍ６（氨基酸序列３８４～５９５）具有红细胞结合作用。Ｍ６的胶体金复合物不能与经胰酶和神经氨酸酶处理过的红细胞发生结合。Ｍ６还能抑制裂殖子入侵红细胞。结论：Ｐ１９５的一段区域（Ｍ６）具有唾液酸残基依赖性红细胞结合作用，它可能是恶性疟原虫裂殖子识别人红细胞的“标志”。Objective:To map out the binding site of P195, which is the major protein on surface of P.falciparum merozoites,to human erythrocytes, and offer a basis for designing malaria vaccine to blockade invasion of merozoites into human erythrocytes. Methods: Eight proteins deriving from P195 were expressed in E.coli, and purified by Ni chelate affinity chromatography. After being refolded, the proteins were labelled with colloidal gold. The colloidal gold protein complexes were incubated with human erythrocytes respectively. In the meantime, the proteins were put into culture supernatant of P.falciparum to observe effects of proteins on invasion of merozoites into erythrocytes. Results: A fragment of P195, M6, whose amino acids sequence is from 384 595, was found to have the ability to bind to human erythrocytes. M6 gold complexes showed no ability to bind to surface of erythrocytes treated by trypsin or neuraminidase. M6 was found to have the ability to inhibit invasion of merozoites of P.falciparum into human erythrocytes. Conclusion: A fragment of P195, M6, has the ability to bind to human erythrocytes. The binding is dependent on sialic acid residues, and may be a basis of recognition of merozoites on erythrocytes.

关 键 词：恶性疟原虫 胶体金 红细胞 P195蛋白 疟原虫 

分 类 号：R382.31[医药卫生—医学寄生虫学]