Purification and characterization of trypsin inhibitor from seeds of faba bean (Vicia faba L.)

Parul Gupta; Kamal Dhawan; Malhotra, S.P.; Randhir Singh

Acta Physiologiae Plantarum 22(4): 433-438

2000


ISSN/ISBN: 0137-5881
DOI: 10.1007/s11738-000-0085-3
Accession: 003538301

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Abstract
A trypsin inhibitor from seeds of faba bean (Vicia faba) was purified to near homogeneity as determined by native-PAGE with about 11% recovery using ammonium sulphate fractionation, ion-exchange chromatography on diethyl amino ethyl-cellulose and gel filtration through Sephadex G-100. The inhibitor had a molecular weight of 18 kD as determined by SDS-PAGE and Sephadex G-100. The inhibitor inhibited trypsin and chymotrypsin up to 50 and 15%, respectively. The inhibition was non-competitive with a dissociation constant in the region of 0.07 mg ml-1 for the enzyme-inhibitor complex. The inhibitor was stable between pH 4 and 5, while activity was completely lost when heated at 125 degrees C for 1 h or at 100 degrees C for 2 h, or when exposed to 2-mercaptoethanol. The faba bean trypsin inhibitor belongs to the Bowman-Birk type of inhibitors, as it has a molecular weight lower than generally observed for Kunitz type inhibitors.

Purification and characterization of trypsin inhibitor from seeds of faba bean (Vicia faba L.)