Oriented Peptide Mixture Libraries can provide a full matrix of preferred and disfavored amino acids at each subsite of an optimal substrate for a new proteinase. This approach is rapid and convenient, requiring only two mixture libraries to complete the analysis. In this paper we demonstrate an extension of this type of analysis, using a focused library employing unnatural amino acids to probe the depth of the S1 position in the catalytic site of the alpha secretase ADAM-10. This analysis indicates that ADAM- 10 will accept amino acids with substantial length and hydrophobicity (e.g. 2- naphthylalanine), but suggests that the S1 site has limitations in the apparent “width” of substituents being presented (e.g. 1-naphthylalanine; gamma branching). A highly selective and efficient substrate for ADAM-10, with a selectivity factor of 380,000 M-1 s -1 , was derived from the predicted consensus substrate. This detailed analysis provides a starting point for the design of inhibitors of this interesting proteinase.
Krstenansky JL, Wang J, Chenail GR, Tiffany MR, Kuesters GM, Natke BC, Nestor Jr JL. Probing proteinase active sites using oriented peptide mixture libraries – ADAM-10. Lett Drug Design Disc. 2004;1:6 –13.