alpha-chymotrypsin and tetrachloroisophthalonitrile

The reaction characteristics of chlorothalonil with glyceraldehyde-3-phosphate dehydrogenase (GPDH), from yeast, (EC 1.2.1.12) were studied in vitro. Enzyme inhibition was related to the amount of [14C]chlorothalonil bound to the protein. Kinetics of enzyme inhibition was non-competitive for the substrate glyceraldehyde-3-phosphate (GAP) (Ki = 0.42 muM). Reversal of enzyme inhibition could not be demonstrated with the low molecular thiol dithiothreitol (DDT), although the thiol did protect the protein against the toxic action of the fungicide. Because 5,5' dithiobis-(2-nitrobenzoic) acid (DTNB) reduced the binding of 14C-labeled fungicide by approximately 90% it is postulated that chlorothalonil affects catalytic activity by reacting with the 4 sulfhydryl sites (cysteine-149) responsible for the binding of GAP. Certain reaction characteristics of the trichloromethyl fulfenyl fungicides with GPDH were found to be similar to those of chlorothalonil. However, chlorothalonil differed from those fungicides in that it did not react with non-thiol groups of either GPDH or alpha-chymotrypsin (alphaCT) and had a slower reaction rate with the GPDH. It is suggested that the differences in reaction rates of the fungicides are due to the molecular size and the chemical nature of the reactive toxiphores.

Topics: Alkylating Agents; Captan; Carbon Radioisotopes; Chlorobenzenes; Chloroform; Chymotrypsin; Dithionitrobenzoic Acid; Dithiothreitol; Electrophoresis, Polyacrylamide Gel; Fungicides, Industrial; Glyceraldehyde; Glyceraldehyde-3-Phosphate Dehydrogenases; Kinetics; Nitriles; Phthalimides; Protein Binding; Sulfhydryl Compounds; Sulfides; Sulfur Radioisotopes