Isolation and characterization of tepary bean proteinase inhibitors and their nutritional and physiological significance.

A protein inhibitor of bovine trypsin and chymotrypsin has been purified from tepary bean (Phaseolus acutifolius) using salt extraction and ammonium sulfate precipitation followed by ion exchange chromatography, hydrophobic chromatography and gel permeation chromatography. Tepary proteinase inhibitor has a molecular weight of 17,500 daltons as measured by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and gel permeation chromatography. Amino acid analysis showed a high content of half-cystine, aspartic acid, serine and glutamic acid residues. The amino-terminal amino acid sequence of the inhibitor revealed high homology to the Bowman-Birk inhibitor family. The tepary proteinase inhibitor has antiproteinase activities similar to those of legume inhibitors of serine proteinase with inhibition of trypsin and chymotrypsin, but not pepsin. Other properties shared with the Bowman-Birk inhibitors include acid and heat resistance, and the presence of two distinct reactive sites, one for trypsin and another for chymotrypsin as indicated by chemical modification of the inhibitor. A similar trypsin and chymotrypsin inhibition was found in a tepary prolamine fraction having a molecular weight of 17,700 daltons estimated by SDS-PAGE and gel permeation chromatography. The amino acid composition of this inhibitor closely resembles those of the low molecular weight Bowman-Birk inhibitors of other legumes. A thermal inactivation study showed that tepary proteinase inhibitor was extremely heat stable at neutral and acidic pH, but heat labile at alkaline pH, similar to the soybean Bowman-Birk and lima bean trypsin inhibitors. Inclusion of tepary bean proteinase inhibitor in diets containing whole egg as the source of dietary protein (8%) significantly reduced growth rate, protein efficiency ratio and protein digestibility, and caused pancreatic hypertrophy in mice