Protease inhibitors from several classes work synergistically against Callosobruchus maculatus.

Targeting multiple digestive proteases may be more effective in insect pest control than inhibition of a single enzyme class. We therefore explored possible interactions of three antimetabolic protease inhibitors fed to cowpea bruchids in artificial diets, using a recombinant soybean cysteine protease inhibitor scN, an aspartic protease inhibitor pepstatin A, and soybean Kunitz trypsin inhibitor KI. scN and pepstatin, inhibiting major digestive cysteine and aspartic proteases, respectively, significantly prolonged the developmental time of cowpea bruchids individually. When combined, the anti-insect effect was synergistic, i.e., the toxicity of the mixture was markedly greater than that of scN or pepstatin alone. KI alone did not impact insect development even at relatively high concentrations, but its anti-insect properties became apparent when acting jointly with scN or scN plus pepstatin. Incubating KI with bruchid midgut extract showed that it was partially degraded. This instability may explain its lack of anti-insect activity. However, this proteolytic degradation was inhibited by scN and/or pepstatin. Protection of KI from proteolysis in the insect digestive tract thus could be the basis for the synergistic effect. These observations support the concept that cowpea bruchid gut proteases play a dual role; digesting protein for nutrient needs and protecting insects by inactivating dietary proteins that may otherwise be toxic. Our results also suggest that transgenic resistance strategies that involve multigene products are likely to have enhanced efficacy and durability.

Lysosomes and protein degradation.

Considerable evidence from studies with group-specific proteinase inhibitors, in particular pepstatin, the aspartic proteinase inhibitor, implicates lysosomes in turnover of endogenous cellular proteins. Recent experiments using a new group-specific inhibitor of thiol (cysteine) proteinases, Z-Phe-Ala-diazomethyl ketone, are described. Lysosomal participation is most clearly established for the degradation of long half-life proteins in situations in which turnover is accelerated because of nutritional or hormonal deficiencies. Some evidence indicating their involvement in 'basal' proteolysis is also discussed. Whether lysosomal proteolysis is selective remains to be established, and possible approaches to this question are outlined.