Identification and molecular characterization of a novel Babesia orientalis thrombospondin-related anonymous protein (BoTRAP1)

作者：Long Yu, Qing Liu, Xueyan Zhan, Yuan Huang, Yali Sun, Zheng Nie, Yangnan Zhao, Xiaomeng An, Muxiao Li, Sen Wang, Yangsiqi Ao, Cuiqin Huang, Lan He, Junlong Zhao · 发表于：Parasites & Vectors · 年份：2018 · DOI：10.1186/s13071-018-3245-2 · 被引用次数：15 · 研究领域：Vector-borne infectious diseases、Invertebrate Immune Response Mechanisms、Leptospirosis research and findings

BACKGROUND: The thrombospondin-related anonymous protein (TRAP) family, a kind of transmembrane protein, is widely distributed with a conserved feature of structure in all apicomplexan parasites and plays a crucial role in the gliding motility and survival of parasites. METHODS: The Babesia orientalis TRAP1 gene (BoTRAP1) was truncated and cloned into a pET-42b expression vector and expressed as a GST-tag fusion protein with a TEV protease site. Rabbit anti-rBoTRAP1 antibody was produced and purified using a protein A chromatography column. Western blot analysis was performed to identify the native protein of BoTRAP1 and differentiate B. orientalis-infected positive from negative serum samples. The localization of BoTRAP1 on merozoites was identified by the indirect florescent antibody test (IFAT). RESULTS: The partial sequence of the TRAP1 gene was cloned from B. orientalis cDNA and identified to contain a von Willebrand factor A (vWFA) region and a thrombospondin type-1 (TSP-1) domain; it had a length of 762 bp, encoding a polypeptide of 254 amino acid residues with a predicted size of 28.2 kDa. The partial sequence was cloned into a pET-42b expression vector and expressed in E. coli as a GST fusion protein. Western blot indicated that rBoTRAP1 has a high immunogenicity and can differentiate B. orientalis-infected positive and negative serum samples collected from water buffaloes. IFAT showed that BoTRAP1 is mainly localized on the apical end of intracellular parasites by u...