Calcium binding to intestinal goblet cell mucin.
1. Binding of Ca-2+ to goblet cell mucin of rat small intestine was studied using equilibrium dialysis against 0.01 M Tris/HCl buffer (pH 7.4) and tracer amounts of 45-CaCl2. Binding was found to reach saturation at a Ca free -2+ concentration of 0.1--1.0 mM, to be independent of temperature (4-37 degrees C), and to increase with increasing pH (5.0-8.7). At low concentrations of Ca free -2+ (smaller than 0.03 mM) the binding curve was sigmoidal, suggesting positive cooperativity of binding sites and a possible change in the tertiary structure of the mucin. Binding was markedly reduced, and sigmoidicity abolished, by removal of sialic acid from the mucin, or by adding 0.14 M NaCl to the dialysis medium. This latter finding suggests that, in vivo, other cations would compete for Ca-2+ binding ligands. 2. Under conditions mimicking those used for binding studies, CaCl2 (10- minus 5 M) was found to cause a small increase (0.03 units) in the absorbance of mucin solutions, especially in the ultraviolet region, possibly indicating increased light scattering. No change in the solubility of the mucin was observed after the addition of CaCl2 (10- minus 6-10- minus 4 M). A significant decrease in viscosity of the mucin was noted, however, with the addition of CaCl2 (10- minus 6-10- minus 2 M). Together with the binding data, these findings suggested that during binding, Ca-2+ combines with negative charges on goblet cell mucin (especially those of sialic acid carboxyl groups) and induces contraction or folding of the macromolecule which promotes cooperative cation binding. No evidence was obtained to suggest that CaCl2 caused precipitation, polymerization or gelation of the mucin in 0.01 M Tris/HCl.?
D000818:Animals; D001665:Binding Sites; D002118:Calcium; D003956:Dialysis; D006863:Hydrogen-Ion Concentration; D007413:Intestinal Mucosa / Q000166:cytology; D007421:Intestine, Small / Q000166:cytology; D007700:Kinetics; D009077:Mucins; D009994:Osmolar Concentration; D011485:Protein Binding; D051381:Rats; D012794:Sialic Acids; D012995:Solubility; D013696:Temperature; D014783:Viscosity
D009077:Mucins; D012794:Sialic Acids; D002118:Calcium