title: Comparative specificity of porcine pancreatic kallikrein and bovine pancreatic trypsin. Importance of interactions N-terminal to the scissible bond.
abstract: Hydrolyses catalyzed by bovine pancreatic trypsin and porcine pancreatic kallikrein were studied using synthetic peptide substrates of the type E chi-L chi 2-L chi 1 decreases Y and E chi-L chi 3-L chi 2-L chi 1 decreases Y with L chi 1 = Arg defining the hydrolysis position (indicated by the arrow). The leaving moiety Y was -OCH3, -NH-C6H4-p-NO2 and -Ala-NH2. Insight into interactions occurring between the active site of the enzymes and the acyl moiety of the substrates was gained by studying the influence on hydrolysis rate of structural variation of residues L chi 2 and L chi 3. Parallel analyses of the hydrolyses of the ester, anilide, and peptide substrates having the same acyl moiety considerably facilitated the interpretation of the kinetic data. Trypsin, but not kallikrein, displayed high reactivity even with relatively short substrates. Ac-Ala-Arg-Ala-NH2, for example, was a better substrate for trypsin than for kallikrein by a factor of 1.3 X 10(4) in terms of kcat and 5.9 X 10(4) in terms of kcat/Km. Reactivity differences of such magnitude were related to two main differences in enzyme-substrate interactions: the interaction of the arginine side chain of the substrate with the specificity pocket of the enzyme is optimal for trypsin but poor for kallikrein and the number of hydrogen bonds formed by the enzyme with the backbone section of the substrate on both sides of the specific residue is larger in the case of trypsin. The latter difference is found to be related to the structure of amino-acid residue 192 which is glutamine in trypsin and methionine in kallikrein.
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002417:Cattle; D006868:Hydrolysis; D007610:Kallikreins / Q000378:metabolism*; D007700:Kinetics; D010179:Pancreas / Q000201:enzymology*; D013379:Substrate Specificity; D013552:Swine; D014357:Trypsin / Q000378:metabolism*
chemical_list: D007610:Kallikreins; D014357:Trypsin