Protease Inhibitors in Phaseolus spp. Seeds

Protease inhibitors (PIs) are widely distributed in plant kingdom and are present in significant amounts in pulse seeds. PIs are grouped in families and superfamilies or clans. The enzyme specificity and inhibitory activity of each inhibitor type depend on the amino acidic residues constituting the reactive binding loop. PIs interfere with the digestive process inhibiting the activity of enzymes such that hydrolyse the proteins into more short peptides. For this reason, they are considered antinutritional compounds and produce adverse effects in insects and animals eating raw seeds. Kunitz trypsin inhibitors (KTIs) and Bowman-Birk inhibitors (BBIs) are the predominant PIs in legume seeds. As consequence of the legume nutritional relevance at worldwide level, both these inhibitors elicited a very high number of studies. BBIs are the prominent PI type present in Phaseolus spp. seeds. They consist of small proteins with molecular mass around 8-9 kDa with approximately 100 amino acids and a high number of cysteine residues. BBIs are double headed inhibitors being able to bind at the same time two identical or different proteinases (trypsin/trypsin, trypsin/chymotrypsin or elastase/trypsin). In common bean (Phaseolus vulgaris L.), BBIs are encoded by a small multigene family composed by three double-headed inhibitors. Genome analysis has showed that all three BBIs clustered at a single locus on chromosome 4. Although, micro-heterogeneity has been described within and among cultivated and wild Phaseolus species, the deduced amino acidic sequences result highly conserved. Moreover, the gene sequences of BBIs has a high similarity with those of other pulses. BBI purified by some Phaseolus species showed high stability to heat and maintenance of activity over a broad range of pH. Conversely, the inhibitory activity of whole seed is significantly reduced (over 80%) by cooking. Otherwise, the BBI content shows a high variation within and among wild and cultivated Phaseolus spp. with higher values mainly associated to the wild species. Drought stress suffered by plant during the vegetative growth stage produces an appreciable increase of BBI level in common bean seeds