MOLECULAR-BASIS OF HUMAN BAND-3 MUTATION ASSOCIATED WITH INCREASED ANION TRANSPORT

Band 3, the anion transporter, is a ubiquitous protein. It is present in brain and all other tissues. Not only is band 3 present in cell mem branes, but also in nuclear, Golgi, and mitochondrion membranes. Band 3 is involved in respiration, acid-base balance, and is the major stru ctural protein linking the plasma membrane to the cytoskeleton. Thus, alterations/mutations in the transport segment of the band-3 molecule might be expected to be of major importance. We discovered and sequenc ed a mutation of band 3, high-transport band 3 (HTbd3), that exhibits anion transport that is 2-3 times above normal. Anion transport studie s of the family members revealed that red cells from the proposita, on e of two siblings, and both parents had abnormally increased anion tra nsport (increased V-max). We used synthetic peptides of band 3 to help localize the change along the band-3 molecule. Results suggest that h igh-transport band 3 is altered in or near residue 869-883. This place s the alteration toward the carboxyl terminal of band 3, cDNA sequenci ng demonstrated that the mutation was a proline to leucine at residue 868. A peptide was synthesized corresponding to residues 853-870 for t esting in the anion transport inhibition assay. This peptide significa ntly inhibited anion transport (p less than or equal to 0.001) indicat ing that it is an anion transport/binding region of band 3. Thus, DNA technology confirms the validity of the anion transport inhibition ass ay for localizing transport regions. Glucose transport is decreased in affected individuals. The HTbd3 mutation appears benign as determined by the red cell aging panel. IgG binding, creatinine, and glyceraldeh yde-3-phosphate dehydrogenase are normal. Our studies indicate that th e most rapid and sensitive techniques for detecting band-3 alterations are polyacrylamide gel electrophoresis, IgG binding, and anion transp ort studies. This is the only mutation of band 3 discovered to date th at affects the transmembrane, anion transport region of band 3.