Inactivation of thermolysin with N-chloroacetyl-N-hydroxy-β-L-phenylalanine N-methylamide

N-Chloroacetyl-N-hydroxy-beta-Phe-NHMe and N-chloroacetyl-N-hydroxy-alpha-Phe-OMe were designed, synthesized and evaluated as irreversible inhibitors of thermolysin, a representative zinc protease. Analysis of kinetic data of the enzymic activity of thermolysin in the presence of these inhibitors revealed that they are indeed potent inactivators of thermolysin having the k(inact)/K-I values of 3.06 and 0.05 M-1 s(-1), respectively. We have established that the inhibitory activity of N-chloroacetyl-N-hydroxy-beta-Phe-NHMe stems mainly from the (R)enantiomer that belongs to the "L" series. The (R)-enantiomer is also responsible for the inactivation in the case of N-chloroacetyl-N-hydroxy-alpha-Phe-OMe, but this enantiomer belongs to the "D"-series.