Involvement of SDS-Stable Higher M_r Forms of Bovine Normal Milk α-Lactalbumin in Inducing Intestinal IEC-6 Cell Death

Abstract

Monomeric 14-kDa bovine α-lactalbumin was purified with a preparation of lower molecular weight whey protein concentrate from Holstein cow normal milk followed by size exclusion chromatography. The protein showed a stimulatory rather than an inhibitory effect on the proliferation of a cultured IEC-6 cell line from the rat small intestine. But incubation in 30% trifluoroethanol/acetate buffer (pH 5.5) at 37 °C for 5 d in a slowly rotating test tube rendered it highly cytotoxic with concomitant appearance of SDS-stable 20- and 30-kDa forms of α-lactalbumin on electrophoresis. Furthermore, α-lactalbumin obtained by a one-step purification procedure by affinity chromatography on an anti-α-lactalbumin antibody column from the lower molecular weight whey protein concentrate, which had been found to contain several SDS-stable higher M_r forms of α-lactalbumin, exhibited potent inhibitory activity on IEC-6 cell growth. These results indicate the involvement of SDS-stable higher M_r forms of bovine normal milk α-lactalbumin in inducing cell death on the intestinal IEC-6 cell line.