Abstract
The nature of bilirubin binding to serum albumin is of considerable interest in the understanding of neonatal hyperbilirubinemia. Each albumin molecule has been shoun to contain 1 high-affinity and 2 low-affinity bilirubin binding sites. The structural analysis of the high-affinity binding site uas undertaken using the technique of affinity labeling. The label uas prepared by reacting bilirubin with Uooduard's reagent K, uhich converted both carboxyl groups of bilirubin into reactive enol esters. Coupling of this bilirubin derivative to albumin was achieved under N2 at pH 9.4. A yellou monomer of albumin uas isolated by gel chromatography. Dialysis of this monomer under denaturing conditions showed covalent attachment of the pigment to the protein. The specificity of the labeling reaction was confirmed by 6 independent methods. Monomer albumin (584 amino acid residues), thus labeled uith 14C bilirubin, was fragmented into 7 peptides by cyanogen bromide cleavage, followed by reduction and carboxymethylation. The bilirubin label uas found to be covalently bound to peptide 3 (residues 124-297) and peptide 6 (residues 447-547). Further structural analysis of this bilirubin-binding site is now in progress.
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Gitzelmann-Cumarasamy, N., Kuenzle, C. & Duc, G. 20: Mapping of the High-Affinity Bilirubin-Binding Site of Human Serum Albumin. Pediatr Res 10, 876 (1976). https://doi.org/10.1203/00006450-197610000-00025
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DOI: https://doi.org/10.1203/00006450-197610000-00025