Screening of compounds interacting with HIV-1 proteinase using optical biosensor technology
1998 (English)In: Analytical Biochemistry, ISSN 0003-2697, E-ISSN 1096-0309, Vol. 265, no 2, 340-350 p.Article in journal (Refereed) Published
A high-resolution optical biosensor assay for screening of low-molecular-weight compounds, using an immobilized protein target, has been developed. HIV-1 proteinase was immobilized on the sensor surface by direct amine coupling and a variety of inhibitors and noninteracting reference drugs were applied to the sensor surface in a continuous how of buffer. The procedure did not require intrinsic reporter groups, substrates, inhibitors, or other ligands for detection. By using a reference protein, the signal could be corrected for the relatively large background signal caused by differences in dimethyl sulfoxide concentration between running and sample buffers. Substances binding with high affinity (K-i in nM range) required efficient regeneration of the sensor surface and washing of the injection system between sample cycles to get consistent results. Analysis was simplified by using report points, extracted during both association and dissociation phases, and a simple graphical display of data. The optimized assay could correctly distinguish HIV-1 inhibitors from other compounds in a randomized series, indicate differences in their interaction kinetics, and reveal artifacts due to nonspecific signals, incomplete regeneration, or carryover. The method is expected to be generally applicable to secondary screening of low-molecular-weight compound libraries with proteins as targets.
Place, publisher, year, edition, pages
1998. Vol. 265, no 2, 340-350 p.
HIV, protease, proteinase, inhibitor, screening, interaction, kinetics, biosensor, Biacore, SURFACE-PLASMON RESONANCE, PROTEASE INHIBITORS, AFFINITIES, BINDING
Biochemistry and Molecular Biology
IdentifiersURN: urn:nbn:se:uu:diva-84906DOI: 10.1006/abio.1998.2927ISI: 000077702000021OAI: oai:DiVA.org:uu-84906DiVA: diva2:112814