title: High-affinity binding of Ca2+ to bovine alpha-lactalbumin in the absence and presence of EGTA.
abstract: Literature values for the Kd for Ca2+ in bovine alpha-lactalbumin range over 3 orders of magnitude. There is a difference between two results obtained with EGTA as a metal-ion buffer, partly because different values for the Kd of Ca2+-EGTA were used in the calculations, and a much wider difference between results obtained in the presence and absence of EGTA, which has been attributed to an interaction between EGTA and the protein. Titrations in a flow-dialysis cell showed that Mn2+ competed with Ca2+ for the high-affinity site on the protein, and the results, combined with a Kd for Mn2+ of 2.1 +/- 0.1 microM, which was determined fluorimetrically, gave a Kd for Ca2+ of 1.3 +/- 0.1 nM. When alpha-lactalbumin containing 45Ca2+ was titrated with EGTA in a flow-dialysis cell, and widely accepted metal-chelation data for EGTA were used in the calculations, a Kd for Ca2+ of 1.10 +/- 0.03 nM was obtained. The results from the two methods are so similar as to indicate that the affinity for Ca2+ was unaffected by the presence of EGTA.
mesh_terms: D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002417:Cattle; D003956:Dialysis; D004533:Egtazic Acid / Q000494:pharmacology*; D005026:Ethylene Glycols / Q000494:pharmacology*; D005260:Female; D007700:Kinetics; D007768:Lactalbumin / Q000378:metabolism*; D008345:Manganese / Q000494:pharmacology; D011485:Protein Binding; D013050:Spectrometry, Fluorescence
chemical_list: D005026:Ethylene Glycols; D008345:Manganese; D004533:Egtazic Acid; D007768:Lactalbumin; D002118:Calcium