Role of the charged residues on the substrate specificity of retroviral proteinases  Page description

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Details of project

  
Identifier
35191
Type F
Principal investigator Boross, Péter
Title in Hungarian Töltéssel rendelkező oldalláncok szerepe retrovirális pro-teinázok szubsztrát-specificitásában
Title in English Role of the charged residues on the substrate specificity of retroviral proteinases
Panel Molecular and Structural Biology and Biochemistry
Department or equivalent Department of Biochemistry and Molecular Biology (University of Debrecen)
Participants Fehér, Anita
Kádas, János
Starting date 2001-01-01
Closing date 2005-12-31
Funding (in million HUF) 5.790
FTE (full time equivalent) 0.00
state closed project





 

Final report

  
Results in Hungarian
A HIV-1 életciklusában betöltött szerepe miatt terápiás (AIDS) célponttá vált retrovirális proteáz (PR) vizsgálatával betekintést nyerhetünk az enzim inhibitorokkal szembeni rezisztencia kialakulásának molekuláris mechanizmusáról. A rezisztenciában megjelenő HIV-1 mutánsok szubsztrát-specificitási, stabilitási, gátolhatósági és szerkezeti vizsgálatai mellett részletesen összehasonlítottuk a HIV-1, HTLV-1, MLV és BLV proteázok tulajdonságait. Ezen vizsgálatokhoz kidolgoztunk egy nagy teljesítményű fluoreszcens mérési módszert. Vizsgálatainkat kiterjesztettük egy klinikai kipróbálás alatt álló ígéretes HIV-1 proteáz inhibitornak, valamint szubsztrát-alapú peptid analóg inhibitoroknak vad tipusú és mutáns HIV-1 proteázokkal alkotott komplexeinek szerkezetvizsgálatával. A HFV PR különleges tulajdonságaiért felelős aminosavak feltérképezése céljából mutáns HFV proteázokkal pH-optimum és urea-stabilitási vizsgálatokat végeztünk.
Results in English
As the retroviral protease became a useful therapeutic target of the AIDS due to its essential role in the life cycle of the HIV-1, the molecular mechanism of the raising resitance against HIV-1 protease inhibitors can be revealed by studying of the enzyme. Besides the substrate specificity, stability, inhibitory and structural studies on the mutant forms of HIV-1 protease appearing in the resistance we compared the features of the wild type HTLV-1, MLV and BLV proteases to the HIV-1 protease. We developed a high-throughput fluorescent method for these studies. We extended our studies to structural analysis of the wild type and mutant HIV-1 proteases complexed with a new potent and promising HIV-1 protease inhibitor which is in clinical trial as well as substrate based peptide analog inhibitors. Urea stability and pH-optimum of mutants of HFV PR were measured to map their special features.
Full text http://real.mtak.hu/86/
Decision
Yes





 

List of publications

  
Mahalingam B, Boross P, Wang YF, Louis JM, Fischer CC, Tozser J, Harrison RW, Weber IT: Combining mutations in HIV-1 protease to understand mechanisms of resistance., Proteins: 48(1): 107-16, 2002
Koh Y, Nakata H, Maeda K, Ogata H, Bilcer G, Devasamudram T, Kincaid JF, Boross P, Wang YF, Tie Y, Volarath P, Gaddis L, Harrison RW, Weber IT, Ghosh AK, Mitsuya H: Novel bis-tetrahydrofuranylurethane containing nonpeptidic protease inhibitor (PI) UIC-94017 (TMC114) with potent activity against multi-PI-resistant HIV in vitro, Antimicrobial Agents and Chemotherapy 47(10):3123-9, 2003
Feher A, Boross P, Sperka T, Oroszlan S, Tozser J: Expression of the murine leukemia virus protease in fusion with maltose binding protein in Escherichia coli., Protein Expression and Purification 35 (1) 62-8, 2004
Tie Y, Boross P, Wang YF, Gaddis L, Hussain AK, Leschenko S, Ghosh AK, Louis JM, Harrison RW, Weber IT: High resolution crystal stuctures of HIV-1 protease with a potent non-peptide inhibitor (UIC-94017) active against multi-drug-resistant clinical strains., Journal of Molecular Biology, 338, 341-352, 2004
Mahalingam B, Wang YF, Boross P, Tőzsér J, Louis JM, Harrison RW, Weber IT: Crystal structures of HIV-1 protease V82A and L90M mutants reveal changes in indinavir binding site, European Journal of Biochemistry 271, 1516-1524, 2004
Bagossi P, Kádas J, Miklóssy G, Boross P, Weber IT, Tőzsér J: Development of a microtiter plate fluorescent asay for inhibition studies on the HTLV-1 and HIV-1 proteinases, Journal of Virologial Methods, 119, 87-93, 2004
Kádas J, Weber IT, Bagossi P, Miklóssy G, Boross P, Tőzsér J: Narrow substrate specificity and sensitivity towards ligand binding site mutations of human T-cell leukemia virus proteinase, Journal of Biological Chemistry, 279, 27148-27157, 2004
Tie Y, Boross P, Wang YF, Gaddis L, Liu F,Chen X, Tozser J, Harrison RW, Weber IT: Molecular basis for substrate recognition and drug resistance from 1.1 to 1.6 angstroms resolution crystal structures of HIV-1 protease mutants with substrate analogs., FEBS J. 272(20):5265-77, 2005
Boross P, Tozser J, Bagossi P: Improved purification protocol for wild-type and mutant human foamy virus proteases., Protein Expression and Purification, in press, 2006
Liu F, Boross P, Wang YF, Tozser J, Louis JM, Harrison RW, Weber IT: Kinetic, stability, and structural changes in high-resolution crystal structures of HIV-1 protease with drug-resistant mutations L24I, I50V, and G73S., J. Mol. Biol. 354(4):789-800, 2005
Sperka T, Boross P, Eizert H, Tozser J, Bagossi P: Effect of mutations on the dimer stability and the pH optimum of the, Prot. Eng. Des. Sel. ( in revision), 2006
Kovalevsky YT, Tie Y, Liu F, Boross P, Wang YF, Leshchenko S, Ghosh, A, Weber IT: Susceptibility of an HIV-1 Protease Inhibitor (UIC-94017) to Highly Drug Resistant Mutations D30N, I50V and L90M, J. Med. Chem. 49(4):1379-1387, 2006
Liu F, Kovalevsky AY, Louis JM, Boross P, Wang YF, Harrison RW, Weber IT: Crystal Structure of F53L Mutant of HIV-1 Protease, J. Mol. Biol. (accepted in 2006), 2006
Sperka T, Miklossy G, Tie Y, Bagossi P, Zahuczky G, Boross P, Weber IT, Harrison RW, Tozser J: Bovine leukemia virus protease: Comparison with human T-cell leukemia virus and human immunodeficiency virus proteases., J. Biol. Chem. (Submitted in 2006), 2006
Feher A, Boross P, Sperka T, Miklossy G, Kadas J, Bagossi P, Oroszlan S, Weber IT, Tozser J: Characterization of the murine leukemia virus protease and its comparison with the HIV-1, Journal of General Virology (accepted in 2005), 2006



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