Proteinase inhibitor II from potatoes: isolation and characterization of its protomer components

Abstract

Proteinase inhibitor II, an inhibitor of chymotrypsin and trypsin, is a heat-stable protein with a dimeric molecular weight of 21 000 that is a component of Russet Burbank potato tubers. Four monomeric isoinhibitor species of molecular weight 10 500 comprise inhibitor II and were isolated by chromatography on phosphocellulose in 8 M urea. Upon removal of the urea, each monomeric species dimerized to yield homogeneous dimers. The three major protomer species, called B, C, and D, and their homogeneous dimers were further characterized. They have similar molecular weights and amino acid compositions, and each has an N-terminal alanine residue. Dimers of purified protomers B, C, and D exhibited full cross-reactivities with each other in immunological double-diffusion assays. Reconstituted dimers possess two binding sites for bovine alpha-chymotrypsin, indicating that each monomer possesses one binding site for this enzyme. Significant differences were noted among the reconstituted dimers in their isoelectric points, immunoelectrophoretic mobilities, ion-exchange properties, and their inhibitory reactivities against trypsin. The properties of the inhibitor II dimeric species are similar but not identical to inhibitors IIa and IIb reported from Japanese potatoes (variety "Danshaku-Imo"), indicating the existence of intervarietal, as well as intravarietal, differences among potato tuber inhibitor II isoinhibitors.