Effect of denaturation of alpha-lactalbumin on the formation of BAMLET (bovine alpha-lactalbumin made lethal to tumor cells).
A complex of alpha-lactalbumin with oleic acid, also known as HAMLET/BAMLET (human/bovine alpha-lactalbumin made lethal to tumor cells), causes apoptosis-like death in tumor cells but has little effect on healthy differentiated cells. The aim of this study was to examine whether irreversible denaturation of alpha-lactalbumin is detrimental to the formation and cytotoxicity of BAMLET. Commercial bovine holo alpha-lactalbumin (1-4% w/v) was heated at 80 degrees C for up to 100 min. With an increasing concentration of protein, the denaturation of alpha-lactalbumin proceeded faster, and aggregation became more extensive. Native and sodium dodecyl sulfate polyacrylamide gel electrophoresis showed that a high proportion of the aggregates was linked by disulfide bonds. BAMLET was prepared from native and heat-treated alpha-lactalbumin according to a previously described chromatographic method. Despite the high content of denatured and aggregated alpha-lactalbumin in the heat-treated samples, their conversion into BAMLET was not negatively affected, resulting in BAMLET complexes partly composed of covalently linked aggregates of alpha-lactalbumin. The cytotoxicity of all prepared BAMLET samples was comparable to that of the control sample prepared from native alpha-lactalbumin (LD(50) = 34.6 +/- 2.7 mumol L(-1)). It was concluded that alpha-lactalbumin is not required to be in its native conformation for the conversion into its biologically active BAMLET complex.
Lisková
Kamila
K
Moorepark Food Research Centre, Teagasc, Moorepark, Fermoy, County Cork, Ireland.
Kelly
Alan L
AL
O'Brien
Nora
N
Brodkorb
André
A
D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D002417:Cattle; D045744:Cell Line, Tumor; D006801:Humans; D007768:Lactalbumin / Q000737:chemistry* / Q000494:pharmacology*; D011487:Protein Conformation; D011489:Protein Denaturation; D017510:Protein Folding; D013329:Structure-Activity Relationship
D007768:Lactalbumin