Narrow substrate specificity and sensitivity toward ligand-binding site mutations of human T-cell Leukemia virus type 1 protease.
Human T-cell leukemia virus type 1 (HTLV-1) is associated with a number of human diseases; therefore, its protease is a potential target for chemotherapy. To compare the specificity of HTLV-1 protease with that of human immunodeficiency virus type 1 (HIV-1) protease, oligopeptides representing naturally occurring cleavage sites in various retroviruses were tested. The number of hydrolyzed peptides as well as the specificity constants suggested a substantially broader specificity of the HIV protease. Amino acid residues of HTLV-1 protease substrate-binding sites were replaced by equivalent ones of HIV-1 protease. Most of the single and multiple mutants had altered specificity and a dramatically reduced folding and catalytic capability, suggesting that mutations are not well tolerated in HTLV-1 protease. The catalytically most efficient mutant was that with the flap residues of HIV-1 protease. The inhibition profile of the mutants was also determined for five inhibitors used in clinical practice and inhibitor analogs of HTLV-1 cleavage sites. Except for indinavir, the HIV-1 protease inhibitors did not inhibit wild type and most of the mutant HTLV-1 proteases. The wild type HTLV-1 protease was inhibited by the reduced peptide bond-containing substrate analogs, whereas the mutants showed various degrees of weakened binding capability. Most interesting, the enzyme with HIV-1-like residues in the flap region was the most sensitive to the HIV-1 protease inhibitors and least sensitive to the HTLV-1 protease inhibitors, indicating that the flap plays an important role in defining the specificity differences of retroviral proteases.
Kádas
János
J
Department of Biochemistry and Molecular Biology, Research Center for Molecular Medicine, Medical and Health Science Center, University of Debrecen, H-4012 Debrecen, P. O. Box 6, Hungary.
Weber
Irene T
IT
Bagossi
Péter
P
Miklóssy
Gabriella
G
Boross
Péter
P
Oroszlan
Stephen
S
Tözsér
József
J
D000595:Amino Acid Sequence; D016282:Aspartic Acid Endopeptidases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D001665:Binding Sites; D016333:HIV Protease / Q000235:genetics; D015497:HIV-1 / Q000201:enzymology / Q000235:genetics; D015368:Human T-lymphotropic virus 1 / Q000201:enzymology*; D006801:Humans; D007700:Kinetics; D008024:Ligands; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D019251:Nucleocapsid / Q000378:metabolism; D009842:Oligopeptides / Q000378:metabolism; D011480:Protease Inhibitors / Q000494:pharmacology; D011993:Recombinant Fusion Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D013379:Substrate Specificity
D008024:Ligands; D009842:Oligopeptides; D011480:Protease Inhibitors; D011993:Recombinant Fusion Proteins; D016282:Aspartic Acid Endopeptidases; D016333:HIV Protease; C079711:HTLV-1 protease