Directed molecular evolution approaches for the selection of novel plant proteinase inhibitors.

Achieving plant resistance to insects was one of the first challenges of plant biotechnology. Several different classes of plant proteins have been shown to be insecticidal towards a range of economically important insects pests. Among them there are the inhibitors of proteinases that act on digestive proteins present in the insect gut. It appears however that some insects can overcome the defensive capacities of their host plant by up-regulating a set of "insensitive" proteinases. Recent developments in molecular biology techniques offer the possibility to use in vitro-directed molecular evolution approaches to obtain variants of proteinase inhibitors with new specific activities. Several authors have already reported on the use of combinatorial approaches to select improved plant inhibitors with enhanced activities against insect proteinases. Beyond the identification of novel molecules with insect anti-nutritional properties, the application of in vitro-directed molecular evolution techniques to plant proteinase inhibitors is also relevant owing to the potential anti-tumoral effects of these inhibitors. This review reports on the use of combinatorial strategies to plant proteinase inhibitors. In particular the case of the application of the phage display technologies to a mustard trypsin inhibitor will be discussed.