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Nonconserved charged residues of the AE2 transmembrane domain alter regulation of anion exchange by pH
Author(s) -
STEWART ANDREW K,
KURSCHAT CHRISTINE,
ALPER SETH L
Publication year - 2007
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.21.6.a1281-c
Subject(s) - transmembrane domain , chemistry , intracellular ph , mutagenesis , dids , mutant , amino acid , xenopus , mutation , transmembrane protein , biochemistry , extracellular , conserved sequence , biophysics , peptide sequence , membrane , biology , gene , receptor
We have investigated the molecular basis for regulation of the ubiquitous AE2/SLC4A2 anion exchanger by intracellular pH (pH i ) and extracellular pH (pH o ) through mutagenesis of C‐terminal transmembrane domain (TMD) amino acid residues not conserved in the closely related but pH‐insensitive AE1/SLC4A1. Selected conserved residues were also studied. AE2‐mediated Cl − /Cl − exchange was measured as DIDS‐sensitive 36 Cl − efflux from Xenopus oocytes by varying pH i at constant pH o , and by varying pH o at near‐constant pH i . Mutation of all 26 non‐conserved charged residues of the AE2 TMD yielded functional protein, but mutations of 4 conserved charged residues reduced or abolished function. Wt pH i ‐sensitivity of anion exchange was reduced upon individual mutation of AE2 TMD residues R921, F922, P1077 and R1107, whereas TMD mutants K1153R, R1155K, R1202L displayed enhanced sensitivity to acidic pH i . In addition, individual mutagenesis of non‐conserved AE2 TMD residues E981, K982 and D1075 to the corresponding AE1 residues significantly acid‐shifted pH o ‐sensitivity. These results demonstrate that multiple conserved charged residues are important for basal transport function of AE2. They further suggests that multiple non‐contiguous TMD amino acid residues contribute to regulation of AE2 by pH, and highlight the complexity of the molecular mechanisms by which pH regulates bicarbonate transporters.