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Biochemical Characterization, Specificity and Inhibition Studies of HTLV-1, HTLV-2, and HTLV-3 Proteases.

ABSTRACT: The human T-lymphotropic viruses (HTLVs) are causative agents of severe diseases including adult T-cell leukemia. Similar to human immunodeficiency viruses (HIVs), the viral protease (PR) plays a crucial role in the viral life-cycle via the processing of the viral polyproteins. Thus, it is a potential target of anti-retroviral therapies. In this study, we performed in vitro comparative analysis of human T-cell leukemia virus type 1, 2, and 3 (HTLV-1, -2, and -3) proteases. Amino acid preferences of S4 to S1' subsites were studied by using a series of synthetic oligopeptide substrates representing the natural and modified cleavage site sequences of the proteases. Biochemical characteristics of the different PRs were also determined, including catalytic efficiencies and dependence of activity on pH, temperature, and ionic strength. We investigated the effects of different HIV-1 PR inhibitors (atazanavir, darunavir, DMP-323, indinavir, ritonavir, and saquinavir) on enzyme activities, and inhibitory potentials of IB-268 and IB-269 inhibitors that were previously designed against HTLV-1 PR. Comparative biochemical analysis of HTLV-1, -2, and -3 PRs may help understand the characteristic similarities and differences between these enzymes in order to estimate the potential of the appearance of drug-resistance against specific HTLV-1 PR inhibitors.

SUBMITTER: Kassay N

PROVIDER: S-EPMC7915765 | biostudies-literature | 2021 Feb

REPOSITORIES: biostudies-literature

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Biochemical Characterization, Specificity and Inhibition Studies of HTLV-1, HTLV-2, and HTLV-3 Proteases.

Kassay Norbert N Mótyán János András JA Matúz Krisztina K Golda Mária M Tőzsér József J

Life (Basel, Switzerland) 20210206 2

The human T-lymphotropic viruses (HTLVs) are causative agents of severe diseases including adult T-cell leukemia. Similar to human immunodeficiency viruses (HIVs), the viral protease (PR) plays a crucial role in the viral life-cycle via the processing of the viral polyproteins. Thus, it is a potential target of anti-retroviral therapies. In this study, we performed in vitro comparative analysis of human T-cell leukemia virus type 1, 2, and 3 (HTLV-1, -2, and -3) proteases. Amino acid preferences ...[more]

PMID: 33562087

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Project description:Extracellular proteases of Porphyromonas gingivalis specific for arginyl peptide bonds are considered to be important virulence factors in periodontal disease. In order to determine the number, inter-relationship and kinetic properties of these proteases, extracellular enzymes with this peptide-bond specificity were purified and characterized from P. gingivalis W50. Three forms, which we denote RI, RI-A and RI-B, accounted for all of the activity in the supernatant. All three enzymes contain an alpha chain of approximately 54 kDa with the same N-terminal amino acid sequence. RI is a heterodimer of non-covalently linked alpha and beta chains which migrate to the same position on SDS/PAGE but which can be resolved by 8 M urea/PAGE. RI-A and RI-B are both monomeric, but the molecular mass of RI-B (70-80 kDa) is significantly increased due to post-translational modification with lipopolysaccharide. All forms show absolute specificity for peptide bonds with Arg in the P1 position and are also capable of hydrolysing N-terminal Arg and C-terminal Arg-Arg peptide bonds. Thus they show limited amino- and carboxy-peptidase activity. For the hydrolysis of Nalpha-benzoyl-L-Arg-p-nitroanilide, the pH optimum is 8.0 at 30 degrees C. The Vmax for all three enzymes is controlled by ionization of two residues with apparent pKas at 30 degrees C of 6. 5+/-0.05 and 9.7+/-0.05, and DeltaH values of approximately 29 kJ/mol and approximately 24 kJ/mol in the enzyme-substrate complex. By analogy with papain, the pKa of 6.5 could be ascribed to a Cys and the pKa of 9.7 to a His residue. E-64 [L-trans-epoxysuccinyl-leucylamide-4-(4-guanidino)butane] is a competitive inhibitor of RI, RI-A and RI-B. Based on physical properties and kinetic behaviour, RI-A appears to be analogous to gingipain from P. gingivalis HG66. However the alpha/beta structure of RI differs significantly from that of the high-molecular-mass multimeric complex of gingipain containing four haemagglutinins described by others. Since the genes for RI and high-molecular-mass gingipain are identical, the data indicate that an alternative processing pathway is involved in the formation of RI from the initial precursor. Furthermore, the identical N-termini and enzymic properties of the catalytic component of RI, RI-A and RI-B suggest that the maturation pathway of the RI precursor may also give rise to RI-A and RI-B. The physiological functions of these isoforms and their role in the disease process may become more apparent through examination of their interactions with host proteins.

Dataset Information

Biochemical Characterization, Specificity and Inhibition Studies of HTLV-1, HTLV-2, and HTLV-3 Proteases.

Publications

Biochemical Characterization, Specificity and Inhibition Studies of HTLV-1, HTLV-2, and HTLV-3 Proteases.

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