CHYMOTRYPSIN INHIBITORY CONFORMATION INDUCED BY AMINO-ACID SIDE-CHAINSIDE-CHAIN INTRAMOLECULAR CH PI INTERACTION/

Dipeptide amides H-D-Leu-Phe-NH-R have been found to assume a conforma tion induced by the CH/pi interaction and to inhibit chymotrypsin stro ngly. A series of benzyl amide derivatives H-D-Leu-Phe-NH-[CH2](n)-C6H 5 (n = 0-4) have been assayed for chymotrypsin, They inhibit the enzym e in a competitive manner and the highest inhibition is achieved by th e amide of n = 1 (K-1 3.6 x 10(-6) M), The activity enhancement is dep endent upon the length of methylene chain, not upon the increase in mo lecular hydrophobicity, indicating the presence of an optimal distance between dipeptide backbone and C-terminal phenyl group for chymotryps in inhibition, The C-terminal phenyl group has been found to interact with chymotrypsin stereospecifically, The R-isomer of H-D-Leu-Phe-NH-C H(CH3)-C6H5 is as active as the benzyl amide, while the S-isomer is ab out twenty-fold less active, When the fluorine atom is introduced at a para-position of the C-terminal phenyl group, the resulting dipeptide H-D-Leu-Phe-NH-CH2-C6H4F-p exhibits about six-times increased inhibit ory activity (K-1 = 6.1 x 10(-7) M; this dipeptide is one of the most potent chymotrypsin inhibitors to date), H-1 NMR conformational analys es of these dipeptide amide derivatives show the CH/pi: interaction be tween D-Leu-isobutyl and Phe-phenyl as a key structural element for ch ymotrypsin inhibition, These structural examinations strongly suggest that in the inhibitory conformation the C-terminal phenyl group fits t he chymotrypsin S-1 site, while the hydrophobic core constructed by D- Leu-Phe CH/pi interaction fits the chymotrypsin S-2 or S-1' site.