Comparative bioinformatic and structural analyses of pepsin and renin.

Pepsin, the archetypal pepsin-like aspartic protease, is irreversibly denatured when exposed to neutral pH conditions whereas renin, a structural homologue of pepsin, is fully stable and optimally active in the same conditions despite sharing highly similar enzyme architecture. To gain insight into the structural determinants of differential aspartic protease pH stability, the present study used comparative bioinformatic and structural analyses. In pepsin, an abundance of polar and aspartic acid residues were identified, a common trait with other acid-stable enzymes. Conversely, renin was shown to have increased levels of basic amino acids. In both pepsin and renin, the solvent exposure of these charged groups was high. Having similar overall acidic residue content, the solvent-exposed basic residues may allow for extensive salt bridge formation in renin, whereas in pepsin, these residues are protonated and serve to form stabilizing hydrogen bonds at low pH. Relative differences in structure and sequence in the turn and joint regions of the β-barrel and ψ-loop in both the N- and C-terminal lobes were identified as regions of interest in defining divergent pH stability. Compared to the structural rigidity of renin, pepsin has more instability associated with the N-terminus, specifically the B/C connector. By contrast, renin exhibits greater C-terminal instability in turn and connector regions. Overall, flexibility differences in connector regions, and amino acid composition, particularly in turn and joint regions of the β-barrel and ψ-loops, likely play defining roles in determining pH stability for renin and pepsin.