Red blood cell band 3: lysine 539 and lysine 851 react with the same H₂dids (4,4′-diisothiocyanodihydrostilbene-2,2′-disulfonic acid) molecule

The band 3 protein of the red blood cell membrane catalyzes anion exchange that is inhibited by the stilbenedisulfonate derivative H₂DIDS (4,4′-diisothiocyanodihydrostilbene-2,2′-disulfonic acid). There is one H₂DIDS binding site per 95,000-Da band 3 polypeptide. The single bound H₂DIDS molecule can react covalently with 2 different lysine residues. The 2 lysines that react covalently with H₂DIDS have been localized directly by sequencing fragments of human band 3 from cells labeled with [³H]H₂DIDS. The most rapid covalent reaction is with Lys-539, in agreement with site-directed mutagenesis studies. The slower reaction is with Lys-851, which is known to be the primary site of binding of another anion transport inhibitor, pyridoxal phosphate (Kawano et al., 1988). These results indicate that the protein is folded to bring these 2 residues into close enough proximity to react covalently with the same H₂DIDS molecule. In addition to defining the residues that react with H₂DIDS, these studies have also defined new in situ proteolytic cleavage sites in band 3.