Department of Parasitology, Asahikawa Medical University, Midorigaoka Higashi 2-1, Asahikawa, 078-8510 Hokkaido, Japan.
Exp Parasitol. 2011 Mar;127(3):693-701. doi: 10.1016/j.exppara.2010.11.005. Epub 2010 Nov 21.
Cysteine peptidases have potent activities in the pathogenesis of various parasitic infections, and are considered as targets for chemotherapy and antigens for vaccine. In this study, two cathepsin B-like cysteine peptidases (EmCBP1 and EmCBP2) from Echinococcus multilocularis metacestodes were identified and characterized. Immunoblot analyses demonstrated that EmCBP1 and EmCBP2 were present in excretory/secretory products and extracts of E. multilocularis metacestodes. By immunohistochemistry, EmCBP1 and EmCBP2 were shown to localize to the germinal layer, the brood capsule and the protoscolex. Recombinant EmCBP1 and EmCBP2 expressed in Pichia pastoris, at optimum pH 5.5, exhibited substrate preferences for Z-Phe-Arg-MCA, Z-Val-Val-Arg-MCA, and Z-Leu-Arg-MCA, and low levels of hydrolysis of Z-Arg-Arg-MCA. Furthermore, recombinant enzymes degraded IgG, albumin, type I and IV collagens, and fibronectin. These results suggested that EmCBP1 and EmCBP2 may play key roles in protein digestion for parasites' nutrition and in parasite-host interactions.
半胱氨酸蛋白酶在各种寄生虫感染的发病机制中具有很强的活性，被认为是化疗靶点和疫苗抗原。本研究鉴定并表征了多房棘球蚴原头蚴中的两种组织蛋白酶 B 样半胱氨酸蛋白酶（EmCBP1 和 EmCBP2）。免疫印迹分析表明，EmCBP1 和 EmCBP2 存在于多房棘球蚴原头蚴的排泄/分泌产物和提取物中。通过免疫组织化学分析，EmCBP1 和 EmCBP2 定位于生殖层、包囊和幼体。在毕赤酵母中表达的重组 EmCBP1 和 EmCBP2 在最佳 pH 值 5.5 时，对 Z-Phe-Arg-MCA、Z-Val-Val-Arg-MCA 和 Z-Leu-Arg-MCA 表现出底物偏好，对 Z-Arg-Arg-MCA 的水解水平较低。此外，重组酶降解 IgG、白蛋白、I 型和 IV 型胶原以及纤维连接蛋白。这些结果表明，EmCBP1 和 EmCBP2 可能在寄生虫营养的蛋白质消化和寄生虫-宿主相互作用中发挥关键作用。
