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Kinetic and thermodynamic characterization of HIV-1 protease inhibitors
Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry.
Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry.
2004 (English)In: Journal of Molecular Recognition, ISSN 0952-3499, Vol. 17, no 2, 106-19 p.Article in journal (Refereed) Published
Abstract [en]

Interaction kinetic and thermodynamic analyses provide information beyond that obtained in general inhibition studies, and may contribute to the design of improved inhibitors and increased understanding of molecular interactions. Thus, a biosensor-based method was used to characterize the interactions between HIV-1 protease and seven inhibitors, revealing distinguishing kinetic and thermodynamic characteristics for the inhibitors. Lopinavir had fast association and the highest affinity of the tested compounds, and the interaction kinetics were less temperature-dependent as compared with the other inhibitors. Amprenavir, indinavir and ritonavir showed non-linear temperature dependencies of the kinetics. The free energy, enthalpy and entropy (DeltaG, DeltaH, DeltaS) were determined, and the energetics of complex association (DeltaG(on), DeltaH(on), DeltaS(on)) and dissociation (DeltaG(off), DeltaH(off), DeltaS(off)) were resolved. In general, the energetics for the studied inhibitors was in the same range, with the negative free energy change (DeltaG < 0) due primarily to increased entropy (DeltaS > 0). Thus, the driving force of the interaction was increased degrees of freedom in the system (entropy) rather than the formation of bonds between the enzyme and inhibitor (enthalpy). Although the DeltaG(on) and DeltaG(off) were in the same range for all inhibitors, the enthalpy and entropy terms contributed differently to association and dissociation, distinguishing these phases energetically. Dissociation was accompanied by positive enthalpy (DeltaH(off) > 0) and negative entropy (DeltaS(off) < 0) changes, whereas association for all inhibitors except lopinavir had positive entropy changes (DeltaS(on) > 0), demonstrating unique energetic characteristics for lopinavir. This study indicates that this type of data will be useful for the characterization of target-ligand interactions and the development of new inhibitors of HIV-1 protease

Place, publisher, year, edition, pages
2004. Vol. 17, no 2, 106-19 p.
Keyword [en]
HIV Protease Inhibitors/*chemistry, HIV-1, Indinavir/chemistry, Kinetics, Pyrimidinones/chemistry, Research Support; Non-U.S. Gov't, Ritonavir/chemistry, Sulfonamides/chemistry, Thermodynamics
National Category
Biochemistry and Molecular Biology
Identifiers
URN: urn:nbn:se:uu:diva-90171PubMedID: 15027031OAI: oai:DiVA.org:uu-90171DiVA: diva2:162433
Available from: 2003-04-01 Created: 2003-04-01 Last updated: 2010-03-09Bibliographically approved
In thesis
1. Interaction Characteristics of Viral Protease Targets and Inhibitors: Perspectives for drug discovery and development of model systems
Open this publication in new window or tab >>Interaction Characteristics of Viral Protease Targets and Inhibitors: Perspectives for drug discovery and development of model systems
2003 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Viral proteases are important targets for anti-viral drugs. Discovery of protease inhibitors as anti-viral drugs is aided by an understanding of the interactions between viral protease and inhibitors. This thesis addresses the characterization of protease-inhibitor interactions for application to drug discovery and model system development.

The choice of a relevant target is essential to molecular interaction studies. Therefore, full-length NS3 protein of hepatitis C virus (HCV) was obtained, providing a more relevant target and a better model for the development of HCV protease inhibitors. In addition, resistance to anti-viral drugs, a serious problem in the treatment of AIDS, prompted the investigation of resistant variants of human immunodeficiency virus (HIV) protease.

Drug resistance was initially explored by characterization of the interactions between a series of closely related inhibitors and resistant variants of HIV protease, using an inhibition assay to determine the inhibition dissociation constants (Ki). The relationship between structure, activity and resistance profiles was not clarified, indicating that the effect of structural changes in the inhibitors and the protease are not predictable and must be analyzed case wise. It was proposed that additional kinetic characterization of the interactions was required and a biosensor-based method allowing for determination of affinity, KD, and interaction rate constants, kon and koff, was adopted. The increased physiological relevance of this method was confirmed, and the affinity data have better correlation with cell culture data. In addition, interactions between clinical inhibitors of HIV protease and enzyme variants indicate that increased dissociation rates (koff) are associated with the development of resistance.

Thermodynamic characterization of the interactions between HIV-1 protease and clinically relevant inhibitors revealed distinct energetic characteristics for inhibitors. The resolution of the energetics of association and dissociation identified an inhibitor with unique interaction characteristics and confirmed the validity of using this method for further characterization of molecular interactions.

This work resulted in the development of model systems for the analysis of kinetics, resistance and thermodynamic characteristics of protein-inhibitor interactions. The results give increased understanding of the biomolecular interactions and can be applied to drug discovery.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2003. 49 p.
Series
Comprehensive Summaries of Uppsala Dissertations from the Faculty of Science and Technology, ISSN 1104-232X ; 822
Keyword
Biochemistry, viral proteases, biomolecular interactions, kinetics, thermodynamics, biosensors, drug discovery, Biokemi
National Category
Biochemistry and Molecular Biology
Research subject
Biochemistry
Identifiers
urn:nbn:se:uu:diva-3342 (URN)91-554-5568-9 (ISBN)
Public defence
2003-04-25, lecture hall B42, Uppsala, 10:15 (English)
Opponent
Available from: 2003-04-01 Created: 2003-04-01 Last updated: 2017-05-04Bibliographically approved

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Danielson, U. Helena

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