Purification and Characterization of a Trypsin Inhibitor from Lily Bulb (Lilium lancifolium)

Abstract

As protein protease inhibitors in foodstuffs influence protein nutrition, characterization of these inhibitors is very important for food science and nutrition. Several proteins from lily bulb for foodstuffs showed strong inhibitory activity toward bovine trypsin. A trypsin inhibitor (LTI-I-1) was purified from lily bulb by a method including column chromatographies on CM-Sepharose CL6 B and DEAE-Sepharose CL-6 B and high-pressure liquid chromatography. Purified LTI-I-1 had a molecular weight of 23 000 and pI of 4.7. The stoichiometry was concluded to be a 1 : 1 molar ratio on the basis of the molecular weight of 24 000 for the trypsin and 23 000 for the inhibitor. The inhibitor contained many aspartic acid residues and 4 half-cystines in its constituent amino acids, and also contained 2 disulfide bridges in its structure. The N-terminal 25 residues of LTI-I-1 were sequenced. The inhibitor was stable in a wide pH range under 50°C and was unstable at the higher temperatures than 80°C. The reactive amino acid of LTI-I-1 was arginine. LTI-I-1 inhibited a-chymotrypsin weakly but did not inhibit elastase and subtilisin. From these results, the inhibitor LTI-I-1 is assumed to belong to the Kunitz soybean inhibitor family.