Screening of compounds interacting with HIV-1 proteinase using optical biosensor technology

A high-resolution optical biosensor assay for screening of low-molecular-we ight compounds, using an immobilized protein target, has been developed. HI V-1 proteinase was immobilized on the sensor surface by direct amine coupli ng and a variety of inhibitors and noninteracting reference drugs were appl ied to the sensor surface in a continuous how of buffer. The procedure did not require intrinsic reporter groups, substrates, inhibitors, or other lig ands for detection. By using a reference protein, the signal could be corre cted for the relatively large background signal caused by differences in di methyl sulfoxide concentration between running and sample buffers. Substanc es binding with high affinity (K-i in nM range) required efficient regenera tion of the sensor surface and washing of the injection system between samp le cycles to get consistent results. Analysis was simplified by using repor t points, extracted during both association and dissociation phases, and a simple graphical display of data. The optimized assay could correctly disti nguish HIV-1 inhibitors from other compounds in a randomized series, indica te differences in their interaction kinetics, and reveal artifacts due to n onspecific signals, incomplete regeneration, or carryover. The method is ex pected to be generally applicable to secondary screening of low-molecular-w eight compound libraries with proteins as targets. (C) 1998 Academic Press.