Nonspecific Electrostatic Binding of Substrates and Inhibitors to Porcine Pepsin
Porcine pepsin has long been recognized to have primary and secondary specificity for hydrophobic amino acids, and the binding of low molecular weight inhibitors (e.g., aliphatic alcohols) is also dominated by hydrophobic interactions1. However, the long standing ‘hydrophobic dogma’ in pepsin catalysis was seriously challenged when Pohl and Dunn2 described a series of polycationic oligopeptides that are among the most reactive synthetic substrates known. These highly hydrophilic molecules should not be well accommodated in a hydrophobic active site. Moreover, the steady state kinetic parameters were remarkably sensitive to the acidity of the medium. As the pH was increased from pH 3 to 6, the specificity number kcat/Km increased by three to four orders of magnitude and approached values close to the diffusion limit. The catalytic turnover number kcat was much less affected by the increase in pH, and the Michaelis constant Km decreased accordingly, by three to four orders of magnitude. A typical pH profile of steady state kinetic parameters, for the substrate Lys-Lys-Ala-Lys-Phe-Phe(NO2)-Arg-Leu, is shown in Figure 1.
KeywordsInhibition Constant Aspartic Proteinase Free Energy Profile Porcine Pepsin Carboxylate Residue
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