| Abstract: | The inactivation of chymotrypsin by 5-benzyl-6-chloro-2-pyrone has been studied. Chloride analysis of the inactivated enzyme suggests that chlorine is no longer present in the complex. 13C NMR spectroscopy of chymotrypsin inactivated with 5-benzyl-6-chloro-2-pyrone-2,6-13 C2 shows the presence of two new resonances from the protein-bound inactivator. The chemical shift values of these resonances are consistent with an intact pyrone ring on the enzyme as well as the replacement of the C-6 chlorine by a different heteroatom. X-ray diffraction analysis at 1.5-A resolution of the inactivator-enzyme complex demonstrates that the gamma-oxygen of the active site serine residue (serine 195) is covalently attached to C-6 of the inactivator and that the pyrone ring is intact. The 5-benzyl group of the inactivator is bound to the enzyme in the hydrophobic specificity pocket. The conformational changes that occur in the protein as a result of complexation with the inactivator are discussed. |
