42. Self-Assembled Small-Molecule Microarrays for Protease Screening and Profiling
    H. D. Urbina, F. Debaene, B. Jost, C. Bole-Feysot, D. E. Mason, P. Kuzmic, J. L. Harris, N. Winssinger
    ChemBioChem 2006, 7, 1790-1797

From libraries to arrays. Peptide nucleic acid encoded libraries can be converted into organized microarrays by hybridization to readily available DNA arrays (see images). Here this method has been applied to the discovery of specific inhibitors for cathepsin K and cathepsin F.

Small-molecule microarrays are attractive for chemical biology as they permit the analysis of hundreds to thousands of interactions in a highly miniaturized format. Methods to prepare small-molecule microarrays from combinatorial libraries by a self-assembly process based on the sequence-specific hybridization of peptide nucleic acid (PNA) encoded libraries to oligonucleotide arrays are presented. A systematic study of the dynamic range for multiple detection agents, including direct fluorescence of attached fluorescein and cyanine-3 dyes, antibody-mediated fluorescence amplification, and biotin-gold nanoparticle detection, demonstrated that individual PNA-encoded probes can be detected to concentrations of 10 pm on the oligonucleotide microarrays. Furthermore, a new method for parallel processing of biological samples by using gel-based separation of probes is presented. The methods presented in this report are exemplified through profiling two closely related cysteine proteases, cathepsin K and cathepsin F, across a 625-member PNA-encoded tetrapeptide acrylate library. A series of the specific cathepsin K and F inhibitors identified from the library were kinetically characterized and shown to correlate with the observed microarray profile, thus validating the described methods. Importantly, it was shown that this method could be used to obtain orthogonal inhibitors that displayed greater than tenfold selectivity for these closely related cathepsins.

DOI : 10.1002/cbic.200600242 

archive ouverte unige:24505