Studies on the substrate specificity of the proteinase of equine infectious anemia virus using oligopeptide substrates.
Abstract: The proteinase of the equine infectious anemia virus (EIAV), a lentivirus closely related to human immunodeficiency virus (HIV), was purified from concentrated virus. The specificity of the enzyme was characterized using oligopeptides representing naturally occurring cleavage sites in the Gag and Gag-Pol polyproteins. The length of the substrate binding pocket was found to be 1-2 residues longer than that of HIV proteinases. Although the EIAV and HIV proteinases cleaved most of the peptides at the same bond, some were hydrolyzed by only the EIAV enzyme. Oligopeptides representing cleavage sites in the nucleocapsid protein were also found to be substrates of the EIAV proteinase. However, these peptides were not hydrolyzed by the HIV proteinases. While peptides representing the corresponding sequences in the first cysteine arrays of the nucleocapsid proteins of HIV-1 and HIV-2 were substrates of the proteinases, peptides representing the homologous sequences in the second Cys arrays were resistant against the proteolytic attack. A three-dimensional model of the EIAV proteinase built on the basis of homology with HIV-1 proteinase was used to interpret the differences. In addition to the oligopeptides representing cleavage sites in the Gag and Gag-Pol polyproteins, the EIAV proteinase was also able to cleave an oligopeptide mimicking a cleavage site in the transmembrane protein. Our results suggest that the specificity of lentiviral proteinases share common characteristics, although substantial differences may exist in hydrolysis of some peptides.
Publication Date: 1993-04-06 PubMed ID: 8384879DOI: 10.1021/bi00064a018Google Scholar: Lookup
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- Journal Article
- Research Support
- Non-U.S. Gov't
- Research Support
- U.S. Gov't
- P.H.S.
Summary
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The study investigates the substrate specificity of the equine infectious anemia virus (EIAV) proteinase, identifying unique characteristics compared to similar enzymes in human immunodeficiency virus (HIV). Specifically, the research presents insights into the way EIAV proteinase interacts with various peptide sequences and its ability to cleave specific peptides.
Method and Experimental Approach
- The proteinase from EIAV was purified from concentrated virus samples. This enzyme is similar to the proteinase found in HIV, making comparisons and contrasts between the two of primary interest.
- Researchers used oligopeptides representing naturally occurring cleavage sites within the Gag and Gag-Pol polyproteins to characterize and observe enzyme activities.
Primary Findings
- The substrate binding pocket (the location of an enzyme where a reaction takes place) of EIAV proteinase was found to be slightly longer than that of HIV proteinases. Length differences could account for varied enzyme behavior.
- Although HIV and EIAV proteinases were able to cleave most peptides at the same bond, some peptides were exclusively cleaved by EIAV enzyme.
- Oligopeptides representing cleavage sites within the nucleocapsid protein acted as substrates, reactive molecules, for the EIAV proteinase but not for HIV proteinases.
- Peptides mimicking sequences in the first cysteine arrays of the nucleocapsid proteins of HIV-1 and HIV-2 were susceptible to cleavage by both enzymes, whereas sequences in the second Cys arrays were resistant.
- The team used a three-dimensional model of the EIAV proteinase, built based on its homology with HIV-1 proteinase, to interpret differences in enzyme function and substrate susceptibility.
- The EIAV proteinase could cleave an oligopeptide mimicking a cleavage site in the transmembrane protein, showcasing the enzyme’s varied substrate specificity.
Implications
- The findings led to insights showing that the specificity of lentiviral proteinases, a group that includes both EIAV and HIV, shares common characteristics.
- The study revealed significant differences in the hydrolysis of some peptides between HIV and EIAV, which could have implications in understanding the virus replication process and may inform future therapeutic strategies.
Cite This Article
APA
Tözsér J, Friedman D, Weber IT, Bláha I, Oroszlan S.
(1993).
Studies on the substrate specificity of the proteinase of equine infectious anemia virus using oligopeptide substrates.
Biochemistry, 32(13), 3347-3353.
https://doi.org/10.1021/bi00064a018 Publication
Researcher Affiliations
- Laboratory of Molecular Virology and Carcinogenesis, National Cancer Institute-Frederick Cancer Research and Development Center, Maryland 21702-1201.
MeSH Terms
- Amino Acid Sequence
- Endopeptidases / isolation & purification
- Endopeptidases / metabolism
- HIV Protease / chemistry
- Infectious Anemia Virus, Equine / enzymology
- Molecular Sequence Data
- Oligopeptides / metabolism
- Protein Processing, Post-Translational
- Proteins / metabolism
- Retroviridae Proteins / metabolism
- Substrate Specificity
Grant Funding
- N01-CO-74101 / NCI NIH HHS
Citations
This article has been cited 10 times.- Mótyán JA, Miczi M, Oroszlan S, Tőzsér J. Specificity of the HIV-1 Protease on Substrates Representing the Cleavage Site in the Proximal Zinc-Finger of HIV-1 Nucleocapsid Protein.. Viruses 2021 Jun 8;13(6).
- Weber IT, Wang YF, Harrison RW. HIV Protease: Historical Perspective and Current Research.. Viruses 2021 May 6;13(5).
- Boso G, Tasaki T, Kwon YT, Somia NV. The N-end rule and retroviral infection: no effect on integrase.. Virol J 2013 Jul 13;10:233.
- Tözsér J. Comparative studies on retroviral proteases: substrate specificity.. Viruses 2010 Jan;2(1):147-165.
- Austin BP, Tözsér J, Bagossi P, Tropea JE, Waugh DS. The substrate specificity of Metarhizium anisopliae and Bos taurus carboxypeptidases A: insights into their use as tools for the removal of affinity tags.. Protein Expr Purif 2011 May;77(1):53-61.
- Kontijevskis A, Prusis P, Petrovska R, Yahorava S, Mutulis F, Mutule I, Komorowski J, Wikberg JE. A look inside HIV resistance through retroviral protease interaction maps.. PLoS Comput Biol 2007 Mar 9;3(3):e48.
- Bagossi P, Sperka T, Fehér A, Kádas J, Zahuczky G, Miklóssy G, Boross P, Tözsér J. Amino acid preferences for a critical substrate binding subsite of retroviral proteases in type 1 cleavage sites.. J Virol 2005 Apr;79(7):4213-8.
- Yovandich JL, Chertova EN, Kane BP, Gagliardi TD, Bess JW Jr, Sowder RC 2nd, Henderson LE, Gorelick RJ. Alteration of zinc-binding residues of simian immunodeficiency virus p8(NC) results in subtle differences in gag processing and virion maturation associated with degradative loss of mutant NC.. J Virol 2001 Jan;75(1):115-24.
- Gustchina A, Kervinen J, Powell DJ, Zdanov A, Kay J, Wlodawer A. Structure of equine infectious anemia virus proteinase complexed with an inhibitor.. Protein Sci 1996 Aug;5(8):1453-65.
- Nagy K, Young M, Baboonian C, Merson J, Whittle P, Oroszlan S. Antiviral activity of human immunodeficiency virus type 1 protease inhibitors in a single cycle of infection: evidence for a role of protease in the early phase.. J Virol 1994 Feb;68(2):757-65.
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