NOVEL VARIANTS OF BBI CODING GENES FROM THE LEGUME LATHYRUS SATIVUS

Protease inhibitors of the Bowman-Birk family (BBIs) are double-headed inhibitors present in many plant species, particularly in legumes. BBIs have been shown to be capable of preventing or suppressing carcinogenic processes in a wide variety of in vitro and in vivo model systems. The ability of BBIs to inhibit cancer cell proliferation in vitro seems to be related to the presence of an active chymotrypsin inhibitor binding site. Due to their potential utility, the study of inhibitor variants -- differing for protein sequence, inhibition performance and selectivity -- might be beneficial in identifying new possible sources of profitable molecules. With the aim to isolate and characterize new isoforms of genes coding for BBIs, we investigated the genetic structure of these genes in the legume Lathyrus sativus. We isolated a pool of BBI-like genes from L. sativus that, although very similar to each other, show some significant differences. In particular, a comparison between their deduced amino-acidic sequences highlights a putative BBI not yet described in the literature. In order to perform functional analyses, L. sativus BBIs were heterologously expressed in the yeast Pichia pastoris. After purification of recombinant proteins, their inhibitory activity was evaluated, by means of enzymatic assays using specific substrates for different serine proteases. In a following step purified recombinant proteins will be tested in a series of in vitro assays in order to verify their anticancer activity.