Isolation of a 5,300-dalton peptide containing a pyridoxal phosphate binding site from the 38,000-dalton domain of band 3 of human erythrocyte membranes.

A hydrophobic 5,300-dalton peptide was isolated from the 38,000-dalton domain of Band 3 by sodium dodecyl sulfate polyacrylamide gel electrophoresis and reversed-phase high-performance liquid chromatography. The peptide was affinity labeled with pyridoxal phosphate and sodium [3H]borohydride when erythrocytes were incubated in vitro. The peptide was not labeled with these agents when cells were incubated in the presence of a specific inhibitor of anion transport, suggesting that the peptide contains at least a part of the active center for the anion transport system in the cell membrane. The peptide was eluted from a reversed-phase high-performance liquid chromatography column with a high concentration of acetonitrile (more than 65%), although the elution pattern of the hydrophobic peptide was not as sharp as that of the soluble peptides. However, a satisfactory separation was achieved when this procedure was employed in combination with sodium dodecyl sulfate polyacrylamide gel electrophoresis.