Physical studies on the rabbit hepatic galactoside-binding protein. Effects of calcium and ligands.
The rabbit hepatic galactoside lectin exhibited noncooperative binding of asialo-orosomucoid in four different nonionic detergents, Triton X-100, octyl glucoside, cetyl eicosaoxyethyleneglycol monoether (Brij 58), and dodecyl octaoxyethyleneglycol monoether (C12E8). The Brij 58-solubilized lectin chromatographed as a single peak upon gel filtration on Sepharose 4B and the molecular weight was determined to be 234,000 in the absence of calcium by sedimentation equilibrium ultracentrifugation. When calcium or calcium plus ligand was added, the molecular weight of the lectin increased to 612,000. A pronounced (15%) decrease in the intrinsic fluorescence of the galactoside lectin was observed upon addition of calcium. Based upon changes in fluorescence, the equilibrium dissociation constant for calcium and binding protein was 1.5 x 10(-3) M. No major changes were detected upon calcium addition by ultraviolet absorption or circular dichroism spectroscopy, nor were there any changes when a ligand such as lactose was added. The number of calcium ions bound, as determined by ultrafiltration, was 3.3 Ca2+/polypeptide chain in the absence of other divalent metals and 1.92 Ca2+/chain in the presence of 100 mM Mg2+. The equilibrium dissociation constant determined in this manner for Ca2+ was 3.5 x 10(-4) M.
D000818:Animals; D001212:Asialoglycoproteins; D002118:Calcium / Q000494:pharmacology*; D002135:Calcium-Binding Proteins; D002352:Carrier Proteins / Q000378:metabolism*; D002633:Chemotaxis; D002942:Circular Dichroism; D003902:Detergents / Q000494:pharmacology*; D007700:Kinetics; D008099:Liver / Q000378:metabolism*; D009004:Monosaccharide Transport Proteins; D009961:Orosomucoid / Q000031:analogs & derivatives / Q000378:metabolism; D033902:Periplasmic Binding Proteins; D011487:Protein Conformation; D011817:Rabbits; D013050:Spectrometry, Fluorescence; D013056:Spectrophotometry, Ultraviolet; D013501:Surface-Active Agents / Q000494:pharmacology*
D001212:Asialoglycoproteins; D002135:Calcium-Binding Proteins; D002352:Carrier Proteins; D003902:Detergents; D009004:Monosaccharide Transport Proteins; D009961:Orosomucoid; D033902:Periplasmic Binding Proteins; D013501:Surface-Active Agents; C017030:asialoorosomucoid; C023550:galactose-binding protein; D002118:Calcium