The sulfhydryl groups of the 35,000-dalton C-terminal segment of band 3 are located in a 9000-dalton fragment produced by chymotrypsin treatment of red cell ghosts.
Five sulfhydryl groups of band 3, the anion-transport protein of the red blood cell membrane, can be labeled by N-ethylmaleimide (NEM). Two of these are located in a 35,000-dalton, C-terminal segment produced by chymotrypsin treatment of cells. Extensive treatment of unsealed ghosts with chymotrypsin results in the disappearance of the 35,000-dalton segment, but its two NEM-binding sites area preserved in a 9000-dalton peptide. The latter must therefore be a proteolytic product of the larger segment. Labeling of sulfhydryl groups of band 3 by an impermeant analog of NEM occurs in inside-out, but not in right-side-out vesicles derived from red cell ghosts, supporting the conclusion that NEM-reactive sulfhydryl groups, including those in the 35,000- and 9000-dalton segments, are exposed at the cytoplasmic face of the membrane. These findings support the conclusion that the 35,000-dalton segment crosses the bilayer, and suggest that the 9000-dalton segment may be a membrane-crossing portion of the 35,000-dalton segment.
D001457:Anion Exchange Protein 1, Erythrocyte; D001798:Blood Proteins; D002918:Chymotrypsin / Q000378:metabolism*; D004910:Erythrocyte Membrane / Q000032:analysis; D006801:Humans; D008970:Molecular Weight; D010446:Peptide Fragments / Q000032:analysis; D013438:Sulfhydryl Compounds / Q000032:analysis*
D001457:Anion Exchange Protein 1, Erythrocyte; D001798:Blood Proteins; D010446:Peptide Fragments; D013438:Sulfhydryl Compounds; D002918:Chymotrypsin