title: Functional reassembly of the anion transport domain of human red cell band 3 (AE1) from multiple and non-complementary fragments.
abstract: We constructed cDNA clones encoding N-terminal, C-terminal and internal polypeptide fragments of the human red cell anion exchanger (band 3; AE1). The internal fragments comprised between one and seven putative transmembrane spans with two or more spans deleted from both termini of the membrane domain of band 3. Sets of three, four or five complementary fragments, which together represented the complete amino acid sequence of the membrane domain, were co-expressed in Xenopus oocytes. Stilbene disulphonate-sensitive chloride uptake assays revealed that all six of the three-fragment combinations and two of the four-fragment combinations reassembled functionally in vivo. Unexpectedly, co-expression of a non-complementary pair of fragments comprising the first five and last seven putative transmembrane spans (i.e. entirely lacking spans six and seven) was also found to be sufficient to generate stilbene disulphonate-sensitive chloride uptake.
authors.affiliation: Department of Biochemistry, School of Medical Sciences, University of Bristol, UK. j.d.groves@bristol.ac.uk
authors.forename: J D
authors.initials: JD
authors.lastname: Groves
authors.forename: L
authors.initials: L
authors.lastname: Wang
authors.forename: M J
authors.initials: MJ
authors.lastname: Tanner
mesh_terms: D000818:Animals; D001457:Anion Exchange Protein 1, Erythrocyte / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D001665:Binding Sites; D003001:Cloning, Molecular; D018076:DNA, Complementary; D004910:Erythrocyte Membrane / Q000378:metabolism; D005260:Female; D006801:Humans; D008958:Models, Molecular; D009865:Oocytes; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism; D016133:Polymerase Chain Reaction; D014176:Protein Biosynthesis; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000378:metabolism; D014982:Xenopus laevis
chemical_list: D001457:Anion Exchange Protein 1, Erythrocyte; D018076:DNA, Complementary; D010446:Peptide Fragments; D011994:Recombinant Proteins