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Cleavage of peptide bonds bearing ionizable amino acids at P1 by serine proteases with hydrophobic S1 pocket |
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| Authors: | Mohammad A. Qasim Jikui Song John L. Markley |
| Affiliation: | a Department of Chemistry, Indiana University Purdue University Fort Wayne, 2101 E. Coliseum Blvd., Fort Wayne, IN 46805, USA b Department of Biochemistry, University of Wisconsin—Madison, WI 53706, USA c Department of Chemistry, Purdue University, West Lafayette, IN 47907, USA |
| Abstract: | Enzymatic hydrolysis of the synthetic substrate succinyl-Ala-Ala-Pro-Xxx-pNA (where Xxx = Leu, Asp or Lys) catalyzed by bovine chymotrypsin (CHYM) or Streptomyces griseus protease B (SGPB) has been studied at different pH values in the pH range 3-11. The pH optima for substrates having Leu, Asp, and Lys have been found to be 7.5-8.0, 5.5-6.0, and ∼10, respectively. At the normally reported pH optimum (pH 7-8) of CHYM and SGPB, the substrate with Leu at the reactive site is more than 25,000-fold more reactive than that with Asp. However, when fully protonated, Asp is nearly as good a substrate as Leu. The pK values of the side chains of Asp and Lys in the hydrophobic S1 pocket of CHYM and SGPB have been calculated from pH-dependent hydrolysis data and have been found to be about 9 for Asp and 7.4 and 9.7 for Lys for CHYM and SGPB, respectively. The results presented in this communication suggest a possible application of CHYM like enzymes in cleaving peptide bonds contributed by acidic amino acids between pH 5 and 6. |
| Keywords: | Chymotrypsin Enzyme kinetic pK shift Serine protease Standard Mechanism inhibitor Streptomyces griseus protease B |
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