Related Experiment Video
Updated: Jul 7, 2026
Purification of Biotinylated Cell Surface Proteins from Rhipicephalus microplus Epithelial Gut Cells
Published on: July 23, 2017
BYC, an atypical aspartic endopeptidase from Rhipicephalus (Boophilus) microplus eggs
Maria Clara L Nascimento-Silva1, Alexandre T Leal, Sirlei Daffre
1Instituto de Bioquímica Médica, Universidade Federal do Rio de Janeiro, Rio de Janeiro, Brazil.
Abstract:
An aspartic endopeptidase was purified in our laboratory from Rhipicephalus (Boophilus) microplus eggs [Logullo, C., Vaz, I.S., Sorgine, M.H., Paiva-Silva, G.O., Faria, F.S., Zingali, R.B., De Lima, M.F., Abreu, L., Oliveira, E.F., Alves, E.W., Masuda, H., Gonzales, J.C., Masuda, A., and Oliveira, P.L., 1998. Isolation of an aspartic proteinase precursor from the egg of a hard tick, Rhipicephalus (Boophilus) microplus. Parasitology 116, 525-532]. Boophilus yolk cathepsin (BYC) was tested as component of a protective vaccine against the tick, inducing a significant immune response in cattle [da Silva, V.I., Jr., Logullo, C., Sorgine, M., Velloso, F.F., Rosa de Lima, M.F., Gonzales, J.C., Masuda, H., Oliveira, P.L., and Masuda, A., 1998. Immunization of bovines with an aspartic proteinase precursor isolated from Rhipicephalus (Boophilus) microplus eggs. Vet. Immunol. Immunopathol. 66, 331-341]. In this work, BYC was cloned and its primary sequence showed high similarity with other aspartic endopeptidases. In spite of this similarity, BYC sequence shows many important differences in relation to other aspartic peptidases, the most important being the lack of the second catalytic Asp residue, considered to be essential for the catalysis of this class of endopeptidases. When we determined BYC cleavage specificity by LC-MS, we found out that it presents a preference for hydrophobic residues in P1 and P1' in accordance to most aspartic endopeptidases. Also, when analyzed by circular dicroism, BYC presented high beta sheet content, also a characteristic of aspartic endopeptidases. On the other hand, although both native and recombinant BYC are catalytically active, they present a very low specific activity, what seems to indicate that this peptidase will digest its natural substrate, vitellin, very slowly. We speculate that such a slow Vn degradative process might constitute an important strategy to preserve egg protein content to the hatching larvae.
Insights
This study investigates Boophilus yolk cathepsin (BYC), an aspartic endopeptidase from tick eggs. Despite unique structural differences, BYC exhibits catalytic activity, suggesting a role in preserving egg nutrients for tick larvae.
Area of Science:
- Biochemistry
- Parasitology
- Veterinary Immunology
Background:
- Rhipicephalus (Boophilus) microplus eggs contain an aspartic endopeptidase, Boophilus yolk cathepsin (BYC).
- BYC has been previously explored as a component in vaccines against ticks, eliciting an immune response in cattle.
Purpose of the Study:
- To clone and characterize the primary sequence of BYC.
- To investigate BYC's catalytic activity, substrate specificity, and structural features.
- To understand BYC's potential role in tick egg development and survival.
Main Methods:
- Cloning of BYC and primary sequence analysis.
- Determination of cleavage specificity using Liquid Chromatography-Mass Spectrometry (LC-MS).
- Analysis of secondary structure content via circular dichroism.
Main Results:
- BYC sequence shows high similarity to other aspartic endopeptidases but lacks the second catalytic Asp residue.
- BYC exhibits preference for hydrophobic residues at P1 and P1' positions, typical of aspartic endopeptidases.
- BYC possesses high beta sheet content, a characteristic structural feature of aspartic endopeptidases.
- Both native and recombinant BYC display catalytic activity but with very low specific activity.
Conclusions:
- BYC's unique structural features, particularly the absence of a key catalytic residue, differentiate it from other aspartic endopeptidases.
- The low catalytic activity of BYC suggests a slow degradation of its natural substrate, vitellin.
- This slow degradation may be a strategy to conserve egg protein content, ensuring nourishment for hatching tick larvae.
