Conversion of α-lactalbumin to a protein inducing apoptosis

Abstract

In this study ␣-lactalbumin was converted from the regular, native state to a folding variant with altered biological function. The folding variant was shown to induce apoptosis in tumor cells and immature cells, but healthy cells were resistant to this effect. Conversion to HAMLET (human ␣-lactalbumin made lethal to tumor cells) required partial unfolding of the protein and a specific fatty acid, C18:1, as a necessary cofactor. Conversion was achieved with ␣-lactalbumin derived from human milk whey and with recombinant protein expressed in Escherichia coli. We thus have identified the folding change and the fatty acid as two key elements that define HAMLET, the apoptosis-inducing functional state of ␣-lactalbumin. Although the environment in the mammary gland favors the native conformation of ␣-lactalbumin that serves as a specifier in the lactose synthase complex, the conditions under which HAMLET was formed resemble those in the stomach of the nursing child. Low pH is known to release Ca 2؉ from the highaffinity Ca 2؉ -binding site and to activate lipases that hydrolyze free fatty acids from milk triglycerides. We propose that this single amino acid polypeptide chain may perform vastly different biological functions depending on its folding state and the in vivo environment. It may be speculated that molecules like HAMLET can aid in lowering the incidence of cancer in breast-fed children by purging of tumor cells from the gut of the neonate.

α-Lactalbumin species variation, HAMLET formation, and tumor cell death

“…In addition, α-lactalbumins form stable folding intermediates when Ca 2+ is released [23]. We have shown that the partially unfolded form of the human protein attains a new function when bound to oleic acid [1]. HAMLET kills tumor cells in vitro and the anti-tumor activity is retained in vivo, in a brain tumor model and in patients with skin papillomas [24,25].…”

“…HAMLET 1 (human α-lactalbumin made lethal to tumor cells) is a complex of apo α -lactalbumin and oleic acid (C18:1) [1] which kills tumor cells while healthy cells are spared. The lethal effect on tumor cells was discovered in a casein fraction, obtained after low pH precipitation of human milk [2].…”

“…Subsequently, MAL was shown to also contain oleic acid, and the conditions required for complex formation were defined by deliberate conversion of native α -lactalbumin to HAMLET in the presence of oleic acid [1]. HAMLET was formed after partial unfolding of α-lactalbumin by EDTA-treatment and ion exchange chromatography on an oleic acid conditioned matrix.…”

BAMLET Activates a Lysosomal Cell Death Program in Cancer Cells

“…Accordingly, new strategies to kill cancer cells by nonapoptotic mechanisms have flourished during the past decade, and many inducers of alternative caspase-independent cell death pathways have been identified (3)(4)(5)(6). Among them is a lipid:protein complex composed of oleic acid and human α-lactalbumin that was originally purified from human milk and termed HAMLET (Human Alpha-lactalbumin Made LEthal to Tumor cells) based on its ability to effectively kill transformed cells while leaving nontransformed cells largely unaffected (7,8). In addition to the broad antitumor activity in vitro, the therapeutic effect of HAMLET has been confirmed in a human glioblastoma xenograft model in rats as well as in human patients with skin papillomas and bladder cancer (9)(10)(11).…”

Introduction: Alpha-Lactalbumin, a Multifunctional Protein that Specifies Lactose Synthesis in the Golgi

“…But the protein also has very interesting properties. In 2000 Svensson and colleagues [12] showed that, under low calcium conditions α-lactalbumin could bind a specific fatty acid, C18:1, and adopt a altered conformation that induced apoptosis in tumor cells but not in normal cells. This altered protein was called HAMLET (human a-lactalbumin made lethal to tumor cells).…”