QAE-Sephadex chromatography of human serum albumin variants

Abstract

Hereditary bisalbuminemia is characterized by the appearance of two albumin bands on cellulose acetate or in agar gel after protein electrophoresis with Tris-barbital buffer at pH 8.8 (2, 8, 12). The variant albumin is inherited codominantly with the normal albumin ( 7, lo), and it may be of either the “fast” or “slow” electrophoretic type (5, 15). Biochemical analysis of albumin variants has generally been limited to the calculation of I$ values based on relative migration distances of the normal and variant albumins on cellulose acetate, agar gel, or starch gel. Purification and partial characterization of three “slow” albumin variants, however, have been reported (3, 6, 16). These authors demonstrated the presence of at least one unique peptide in the “slow” albumin variants they studied. Because albumin variants demonstrate ultracentrifugal and immunochemical properties identical to those of normal albumin (4), other means must be used for isolation and purification of these variants. Since fast and slow albumin variants do differ from normal albumin in net charge at a given pH, it is possible to isolate these proteins by ion-exchange chromatography ( 3, 9). We herein report a new method for the separation and purification of severa human serum albumin variants using QAE-Sephadex’ chromatography and a pH gradient elution technique. Purified fast albumin variants can be rapidly prepared by this technique, and they appear to be suitable for albumin-ligand binding studies and for peptide mapping.