Mapping of the S' subsites of porcine pancreatic and human leucocyte elastases.

Trifluoroacetyl dipeptide anilides have been synthesized and used to map the S' subsites of porcine pancreatic elastase and human leucocyte elastase. A confident mapping of these subsites, at least for the porcine enzyme, was possible since the x-ray crystallographic study of its complex with CF3CO-Lys-Ala-NH-Ph-p-CF3 at a resolution of 2.5 A (Hughes, D. L., Sieker, L. C., Bieth, J., and Dimicoli, J. L. (1982) J. Mol. Biol. 162, 645-658) shows the CF3CO group at the S1 subsite and the dipeptide anilide bound at sites close to the S'1-S'3 subsites. Furthermore the effect of substitution was easy to investigate since these ligands are reversible competitive inhibitors of elastases, whose mode of binding to the porcine enzyme has been shown by nuclear magnetic resonance spectroscopy (Dimicoli, J. L., Renaud, A., and Bieth, J. (1980) Eur. J. Biochem. 107, 423-432) to be essentially unique and common to all CF3CO peptide anilides. The total number of S' subsites was found to be three for both enzymes. The individual subsites have the following specificities: subsite S'1, in porcine pancreatic elastase this subsite prefers Lys to Ala or Glu. In human leucocyte elastase this subsite is less specific; subsite S'2, in porcine pancreatic elastase, this subsite has a marked specificity for Ala. It accommodates bulkier residues with some difficulty. In human leucocyte elastase there is a remarkable specificity for Leu at this subsite; subsite S'3, in porcine pancreatic elastase, this subsite has a high aromatic specificity. In human leucocyte elastase there is no such affinity in S'3 but favorable local interaction exists. These specificities are examined on the basis of the coordinates of the CF3CO-Lys-Ala-NH-Ph-p-CF3 . porcine pancreatic elastase complex. Furthermore the different specificities of the S'2 subsite found in our own work and proposed by Atlas (Atlas, D. (1975) J. Mol. Biol. 93, 39-53) are briefly discussed.