Lee K W, Shalaby K A, Thakur A, Medhat A M, Karim A M, LoVerde P T
Department of Microbiology, School of Medicine and Biomedical Sciences, State University of New York, Buffalo 14214, USA.
Mol Biochem Parasitol. 1995 May;71(2):221-31. doi: 10.1016/0166-6851(95)91598-o.
As molecules on the surface or associated with the outer covering (tegument) of Schistosoma mansoni are a major focus as potential vaccine candidates, affinity purified antibodies which are specific to the tegumental antigens were used to immunoscreen a lambda gt11 S. mansoni cercarial cDNA library. One of the identified clones was found to encode the glycolytic enzyme phosphoglycerate kinase (PGK, EC 2.7.2.3). The 1.5-kb cDNA clone has a single open reading frame encoding 416 amino acids and exhibits over 60% identity to PGKs from a number of eukaryotic species. Recombinant S. mansoni PGK (SmPGK) was overexpressed in Escherichia coli, purified, and shown to have PGK enzyme activity. Native protein affinity purified from S. mansoni adult worms was shown by microsequencing to have the same amino-acid sequence as deduced from the cDNA sequence, thus confirming the cDNA clone we identified encodes S. mansoni phosphoglycerate kinase. Antibodies localize the native SmPGK to various tissues including the tegument of 3-h schistosomula and 42-day adult worms.
由于曼氏血吸虫表面或与外层被膜（皮层）相关的分子是潜在疫苗候选物的主要研究重点，因此使用针对皮层抗原的亲和纯化抗体对λgt11曼氏血吸虫尾蚴cDNA文库进行免疫筛选。所鉴定的一个克隆被发现编码糖酵解酶磷酸甘油酸激酶（PGK，EC 2.7.2.3）。这个1.5kb的cDNA克隆有一个单一的开放阅读框，编码416个氨基酸，与许多真核生物物种的PGK有60%以上的同源性。重组曼氏血吸虫PGK（SmPGK）在大肠杆菌中过量表达、纯化，并显示具有PGK酶活性。通过微量测序表明，从曼氏血吸虫成虫中亲和纯化的天然蛋白与从cDNA序列推导的氨基酸序列相同，从而证实我们鉴定的cDNA克隆编码曼氏血吸虫磷酸甘油酸激酶。抗体将天然SmPGK定位到包括3小时童虫和42日龄成虫皮层在内的各种组织中。
