Structure and ligand-binding properties of abnormal human albumins

Abstract

Albumins Redhill, Warwick-1 and Carlisle are monomeric slow albumin variants discovered in sera obtained from patients in unrelated families resident in the U.K. Albumin Redhill had previously been studied in this laboratory and was here purified by fast protein liquid chromatrography (FPLC) and was subsequently submitted for amino acid sequencing by the solid-phase Edman process. It was found that Albumin Redhill has an extra N-terminal arginine residue, and this places it into a new class of albumin variants. The binding of nickel and copper was studied in greater depth than previously, and the binding of these metal ions at the primary N-terminal site confirmed these as being significantly inhibited due to the inclusion of the extra basic amino acid residue. The binding of warfarin to Albumin Redhill is reduced compared to normal albumin. Albumin Carlisle, a heat-stable variant, was found in three members of a family of English origin from Carlisle. The binding of a range of dyes and the electrophoretic mobility on a series of media were assessed. The variant Albumin Carlisle was purified to homogeneity by FPLC chromatofocusing and was shown to be antigenically indistinguishable from albumin A, although it does have a more basic isoelectric point (5.74 compared to 5.63 for normal albumin). The evidence from both electrophoretic and chromatographic procedures are consistent with an acid + neutral amino acid mutation, and studies of the CNBr fragments of the variant suggest that the site of mutation is in the region 329-548 residues. Reverse-phase HPLC has been used to pinpoint a difference in the profile of the tryptic digest of the variant albumin from the normal, and it may be that this technique could be utilised to obtain molecular data on the mutation. The ligand binding properties of metal ions, bilirubin, palmitate and warfarin were assessed and it was shown that Albumin Carlisle has increased warfarin binding but decreased bilirubin affinity, although the binding of metal ions and palmitate was unaffected