IrCL1-The haemoglobinolytic cathepsin L of the hard tick, Ixodes ricinus
IrCL1-The haemoglobinolytic cathepsin L of the hard tick, Ixodes ricinus
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DOI：
10.1016/j.ijpara.2011.06.006
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发表时间：
2011-10-01
影响因子：
4
通讯作者：
Kopacek, Petr
中科院分区：
文献类型：
--
作者：
Franta, Zdenek;Sojka, Daniel;Kopacek, Petr
蜱类摄取血液蛋白的胞内蛋白水解是一个重要的生理过程。在我们的模式蜱，硬蜱，组织蛋白酶L（IrCL 1）是肠道相关的多肽酶复合物的一部分，其内肽酶活性是重要的血红蛋白溶解的初始阶段。我们提出了这种酶的功能和生化特性。我们表明，通过RNA干扰（RNAi），组织蛋白酶L样活性的峰值在缓慢喂养期的女性与IrCL 1。重组IrCL 1在细菌和酵母中表达。肽基和生理蛋白质底物（血红蛋白和白蛋白）的活性分析表明，IrCL 1是一种酸性肽酶，具有非常低的最适pH（3-4），在pH 5以上不稳定。这表明，通过间接荧光显微镜证实的内/溶酶体定位，免疫定位在消化肠细胞的囊泡内的IrCL 1。通过位置扫描合成组合文库和抑制曲线确定的切割特异性表明，IrCL 1具有半胱氨酸肽酶的组织蛋白酶L亚家族的配体结合特性。当IrCL 1沉默的蜱与对照相比进食能力降低时，证明了非冗余的蛋白水解功能。这些数据表明，IrCL 1可能是一个有前途的目标，对蜱和蜱传病原体。(C)2011年澳大利亚寄生虫学会由爱思唯尔有限公司出版。保留所有权利。
Intracellular proteolysis of ingested blood proteins is a crucial physiological process in ticks. In our model tick, Ixodes ricinus, cathepsin L (IrCL1) is part of a gut-associated multi-peptidase complex; its endopeptidase activity is important in the initial phase of haemoglobinolysis. We present the functional and biochemical characterisation of this enzyme. We show, by RNA interference (RNAi), that cathepsin L-like activity that peaks during the slow feeding period of females is associated with IrCL1. Recombinant IrCL1 was expressed in bacteria and yeast. Activity profiling with both peptidyl and physiological protein substrates (haemoglobin and albumin) revealed that IrCL1 is an acidic peptidase with a very low optimum pH (3-4) being unstable above pH 5. This suggests an endo/lysosomal localisation that was confirmed by indirect fluorescence microscopy that immunolocalised IrCL1 inside the vesicles of digestive gut cells. Cleavage specificity determined by a positional scanning synthetic combinatorial library and inhibition profile indicated that IrCL1 has the ligand-binding characteristics of the cathepsin L subfamily of cysteine peptidases. A non-redundant proteolytic function was demonstrated when IrCL1-silenced ticks had a decreased ability to feed compared with controls. The data suggest that IrCL1 may be a promising target against ticks and tick-borne pathogens. (C) 2011 Australian Society for Parasitology Inc. Published by Elsevier Ltd. All rights reserved.