Functional determinants of the Epstein-Barr virus protease
Abstract
Herpesvirus proteases are essential for the production of progeny virus. They cleave the assembly protein that fills the immature capsid in order to make place for the viral DNA. The recombinant protease of the human gamma-herpesvirus Epstein-Barr virus (EBV) was expressed in Escherichia coli and purified. Circular dichroism indicated that the protein was properly folded with a secondary structure content similar to that of other herpesvirus proteases. Gel filtration and sedimentation analysis indicated a fast monomer-dimer equilibrium of the protease with a Kd of about 60 μM. This value was not influenced by glycerol but was lowered to 1.7 μM in the presence of 0.5 M sodium citrate. We also developed an HPLC-based enzymatic assay using a 20 amino acid residue synthetic peptide substrate derived from one of the viral target sequences for the protease. We found that conditions that stabilised the dimer also led to a higher enzymatic activity. Through sequential deletion of amino acid residues from either side of the cleavage site, the minimal peptide substrate for the protease was determined as P5-P2′. This minimal sequence is shorter than that for other herpesvirus proteases. The implications of our findings are discussed with reference to the viral life-cycle. These results are the first ever published on the EBV protease and represent a first step towards the development of protease inhibitors.
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BibTeX
@article{buisson2026functional,
title = {{Functional determinants of the Epstein-Barr virus protease}},
author = {Marlyse Buisson and Erika Valette and Jean-François Hernandez and Florence Baudin and Christine Ebel and Patrice Morand and Jean-Marie Seigneurin and Gérard Arlaud and Rob W.H. Ruigrok},
journal = {Journal of Molecular Biology},
year = {2026},
month = oct,
doi = {10.1006/jmbi.2001.4854},
url = {https://hal.science/hal-05285326v1},
note = {CC BY-NC-ND 4.0},
}
RIS
TY - JOUR
TI - Functional determinants of the Epstein-Barr virus protease
AU - Marlyse Buisson
AU - Erika Valette
AU - Jean-François Hernandez
AU - Florence Baudin
AU - Christine Ebel
AU - Patrice Morand
AU - Jean-Marie Seigneurin
AU - Gérard Arlaud
AU - Rob W.H. Ruigrok
PY - 2026
DA - 2026/10/30
JO - Journal of Molecular Biology
DO - 10.1006/jmbi.2001.4854
UR - https://hal.science/hal-05285326v1
AB - Herpesvirus proteases are essential for the production of progeny virus. They cleave the assembly protein that fills the immature capsid in order to make place for the viral DNA. The recombinant protease of the human gamma-herpesvirus Epstein-Barr virus (EBV) was expressed in Escherichia coli and purified. Circular dichroism indicated that the protein was properly folded with a secondary structure content similar to that of other herpesvirus proteases. Gel filtration and sedimentation analysis indicated a fast monomer-dimer equilibrium of the protease with a Kd of about 60 μM. This value was not influenced by glycerol but was lowered to 1.7 μM in the presence of 0.5 M sodium citrate. We also developed an HPLC-based enzymatic assay using a 20 amino acid residue synthetic peptide substrate derived from one of the viral target sequences for the protease. We found that conditions that stabilised the dimer also led to a higher enzymatic activity. Through sequential deletion of amino acid residues from either side of the cleavage site, the minimal peptide substrate for the protease was determined as P5-P2′. This minimal sequence is shorter than that for other herpesvirus proteases. The implications of our findings are discussed with reference to the viral life-cycle. These results are the first ever published on the EBV protease and represent a first step towards the development of protease inhibitors.
SN - 0022-2836
SN - 1089-8638
ER -