PURIFICATION AND CHARACTERIZATION OF A DIGESTIVE CYSTEINE PROTEINASE FROM THE FIELD SLUG (DEROCERAS-RETICULATUM) - A POTENTIAL TARGET FOR SLUG CONTROL

Proteinase activities present in crop, digestive gland, and salivary g land extracts of the pest slug species, Deroceras reticulatum (Muller) , were investigated. The digestive gland was found to be responsible f or approximate to 80% of the total proteolytic activity against the pl ant protein rubisco. This activity, also detected by maximal hydrolysi s of the synthetic substrate Z-Phe-Arg-pNA at pH 6.0, was activated by thiol compounds and inhibited by the cysteine-specific proteinase inh ibitors E-64 and chicken egg white cystatin (azocasein as substrate). Furthermore, activity was largely unaffected by class-specific inhibit ors diagnostic for aspartic acid proteinases, metalloproteinases, or s erine proteinases. Similar studies demonstrated that the weak proteoly tic activities in the crop and salivary glands were due to serine prot einases and metalloproteinases. Cation-exchange chromatography of dige stive gland extracts showed that the activity in the digestive gland w as due to a single protein of approximate M-r 40 000, which had kineti c properties similar to those of cathepsin L. Inhibition constants of phytocystatins tested against this purified proteinase ranged between 1.28 x 10(-7) and 6.55 x 10(-7) M. These results suggest that the expr ession of phytocystatins in transgenic plants may be an alternative me thod for controlling slug populations in the field.