CRYSTAL-STRUCTURE OF AN ELASTASE-SPECIFIC INHIBITOR ELAFIN COMPLEXED WITH PORCINE PANCREATIC ELASTASE DETERMINED AT 1.9 ANGSTROM RESOLUTION

The crystal structure of a stoichiometric complex between an elastose- specific inhibitor elafin and porcine pancreatic elastase (PPE) has be en determined and refined to a crystallographic R-factor of 19.7% at 1 .9 Angstrom resolution. The polypeptide chain of elafin has a planar s piral shape with an exposed external part and an internal core part wh ich resembles both the crystal structure of human seminal plasma inhib itor (HUSI-1) [Grutter, M. G., Fendrich, G., Huber, R., & Bode, W, (19 88) EMBO J. 7, 345-351] and the solution structure of Na+,K+-ATPase in hibitor (SPAI-1) revealed by NMR analysis [Kozaki, T., Kawakami, Y., T achibana, S., Hatanaka, H., & Inagaki, F. (1994) Pept. Chem., 405-408] . The external region containing the primary binding loop is interconn ected by four disulfide bonds to the internal part composed of a beta- sheet and a hairpin loop. The scissile peptide bend Ala24i(P1)-Met25i( P1') in the primary binding site is intact, and its carbonyl carbon is in van der Waals contact with O gamma of the active site Ser195 of PP E. The seven residues of Leu20i(P5)-Leu26i(P2') of the primary binding loop and the three residues of Ser48i, Cys49i, and Ala52i of the adja cent hairpin loop are in contact with PPE by hydrogen bonds and/or van der Waals interactions in a manner similar to that observed for other serine protease-inhibitor complexes. Electron densities of the N-term inal residues Alali-Ser10i which are not responsible for the elastase inhibitory activity were not visible, probably due to disordered confo rmation. The guanido group (N eta 1, N eta 2) of Arg61 in the complex interacts with S delta of Met25i(P1') by possible hydrogen bonds betwe en N and S atoms, accompanying a large positional shift of the side ch ain of Arg61 (S1') between the complexed and free forms of PPE. The pr imary binding site is stabilized by hydrogen bonds between the guanido group (N eta 1, N eta 2) of Arg22i(P3) and the carbonyl group of Met2 5i(P1') across the scissile bond, as well as by a hydrogen bond betwee n the amino group of Cys23i(P2) and the carbonyl group of Ser48i in th e internal core. This intramolecular hydrogen bond network and the net work of four disulfide bonds might play a significant role in stabiliz ing the conformation of the binding site for expressing the potent spe cific inhibitory activity.