Purification and properties of an α-amylase tetrameric inhibitor from wheat kernel

Abstract

A family of tetrameric protein inhibitors active against exogenous α-amylases has been extracted from wheat kernel with 150 mM NaCl and purified by gel filtration in the absence and presence of guanidine hydrochloride and by ion-exchange chromatography. The purified isoinhibitor fraction showed four bands of protein with closely related electrophoretic mobilities and isoelectric points. The M r was 48000 in the absence of dissociating agents and dropped to about 12 000 in the presence of guanidine or sodium dodecyl sulphate. Amino acid composition, thermostability and high number of intrachain disulphide bridges were properties closely related to those of monomeric and dimeric protein inhibitors from the same source, but the immunological properties were distinctly different. The tetrameric inhibitor was strongly active against animal α-amylases, and inactive against bacterial, fungal and plant enzymes; the extent of inhibition of yellow mealworm α-amylase was dependent, as for many other wheat protein inhibitors, upon preincubation time, pH and temperature. It is likely that these tetrameric inhibitors, together with wheat inhibitor families of M r 24 000 and 12 000, have a common genetic origin.