Because hypersecretion of gallbladder (GB) mucus occurs in gallstone formation and because binding of Ca2+ to biliary lipids only accounts for 50% of the total Ca2+ in GB bile, we investigated the binding of Ca2+ to human biliary mucin. Biliary mucin was purified from GB bile and binding to Ca2+ studied. Scatchard plot analysis suggested two binding sites. Removal of sialic acid by neuraminidase resulted in 10% reduction of Ca2+ binding, whereas, sulfatase treatment reduced Ca2+ binding by 30%. Using a hypotonic NaCl solution, Ca2+ binding to mucin increased curvilinearly with mucin concentration. However, binding decreased with increasing ionic strength of the NaCl solution. We conclude that binding of Ca2+ to mucin is effected mainly through sulfate. Binding to Ca2+ can be displaced by Na+. Ca2+ binding to mucins is enhanced in the setting of low Na+ concentrations. This phenomenon has pathophysiologic implications for the formation of thick mucus in cystic fibrosis epithelia. (C) 2003 Elsevier Inc. All rights reserved.