Centro de Biotecnologia, Universidade Federal do Rio Grande do Sul, Porto Alegre, Brazil.
Comp Biochem Physiol B Biochem Mol Biol. 2010 Dec;157(4):326-35. doi: 10.1016/j.cbpb.2010.08.002. Epub 2010 Aug 11.
The aim of the present study was to address the involvement of Rhipicephalus microplus larval cysteine endopeptidase (RmLCE) in protein digestion in R. microplus larvae and adult females. In this work, an improved purification protocol for native RmLCE was developed. Partial amino acid sequence of the purified enzyme indicates that it is the same enzyme as Boophilus microplus cathepsin-L1 (BmCL1). When vitellin (Vt) degradation by egg and larval enzymes was analyzed, stage-specific differences for RmLCE activity in comparison to vitellin-degrading cysteine endopeptidase (VTDCE) were observed. RmLCE is also able to degrade host hemoglobin (Hb). In agreement, an acidic cysteine endopeptidase activity was detected in larval gut. It was shown that cysteine and aspartic endopeptidases are involved in Vt and Hb digestion in R. microplus larvae and females. Interestingly, we observed that the aspartic endopeptidase Boophilus yolk cathepsin (BYC) is associated with a cysteine endopeptidase activity, in larvae. Synergic hemoglobin digestion by BYC and RmLCE was observed and indicates the presence of an Hb-degrading enzymatic cascade involving these enzymes. Our results suggest that RmLCE/BmCL1 has a continued role in vitellin and hemoglobin digestion during tick development.
本研究旨在探讨硬蜱幼虫半胱氨酸内肽酶（RmLCE）在硬蜱幼虫和雌蜱蛋白消化中的作用。在这项工作中，开发了一种改进的天然 RmLCE 纯化方案。纯化酶的部分氨基酸序列表明，它与 Boophilus microplus 组织蛋白酶-L1（BmCL1）是同一种酶。当分析卵和幼虫酶对卵黄蛋白（Vt）的降解时，与卵黄蛋白降解半胱氨酸内肽酶（VTDCE）相比，观察到 RmLCE 活性在不同阶段的特异性差异。RmLCE 还能够降解宿主血红蛋白（Hb）。同样，在幼虫肠道中检测到酸性半胱氨酸内肽酶活性。结果表明，半胱氨酸和天冬氨酸内肽酶参与了硬蜱幼虫和雌蜱的 Vt 和 Hb 消化。有趣的是，我们观察到天冬氨酸内肽酶 Boophilus 卵黄组织蛋白酶（BYC）与一种半胱氨酸内肽酶活性相关，在幼虫中。BYC 和 RmLCE 协同血红蛋白消化，表明存在涉及这些酶的血红蛋白降解酶级联反应。我们的结果表明，RmLCE/BmCL1 在蜱的发育过程中继续在卵黄蛋白和血红蛋白消化中发挥作用。
