Revealing favorable and unfavorable residues in cooperative positions in protease cleavage sites.

Proteases play critical roles in a wide variety of fundamental biological functions, and numerous protease inhibitors have been developed to treat various diseases including cancer. A wide range of experimental and computational methods have been developed to investigate the specificity and catalytic mechanisms of proteases. However, these methods only focused on the preferences of a single position around a cleavage site in a substrate, rarely on the compositionality of the subsites. We present new methods to quantify the specificity of proteases by considering the combinatorial patterns of amino acid residuals of cleavage sites in substrates. By incorporating the preference at positions, we modeled three types of favorable combinations of residues in cleavage sites. Moreover, by constructing a relationship weight matrix of residues between two positions, we can easily identify unfavorable combinations of residues at the positions. Applying these methods to a set of known cleavage sites of proteases, we revealed numerous favorable and unfavorable residues in cooperative positions in the protease cleavage sites. The results can help understand the specificity and catalytic mechanisms of proteases. To our knowledge, this is the first study that quantifies unfavorable combinations of amino acids between two sites. Furthermore, this method is not limited to the study of proteases and cleavage sites, and can be generalized to uncover the relationships of residues at meaningful sites in other proteins.