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Structure and role of the first membrane‐spanning domain of the epithelial sodium channel
Author(s) -
Kashlan Ossama B.,
Maarouf Ahmad B.,
Kussius Cassandra,
Blumenthal Kenneth M.,
Kleyman Thomas R.
Publication year - 2006
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.20.4.a329
Subject(s) - epithelial sodium channel , protein subunit , chemistry , extracellular , biophysics , intracellular , mutant , ion channel , sodium , selectivity , sodium channel , crystallography , biochemistry , biology , gene , receptor , organic chemistry , catalysis
Epithelial Na + Channels (ENaC) are comprised of three subunits that have been proposed to be arranged in either an α 2 βγ or a higher ordered configuration. Each subunit has two putative membrane‐spanning domains (M1 and M2), intracellular N‐ and C‐ termini, and a large extracellular loop. We have systematically mutated each M1 residue in the α subunit to Trp, and endogenous M1 Trp’s to Ala in order to experimentally determine the secondary structure of M1, and the role of M1 in ENaC pore formation. Na + current and Li + :Na + selectivity of M1 mutants varied with a periodicity consistent with an α‐helical secondary structure. Throughout M1, changes in channel activity correlated inversely with changes in Li + :Na + selectivity, suggesting changes in ion permeability. Preliminary surface expression data suggests that the changes induced by these mutations are caused by changes in the single channel properties of ENaC. Our results provide the first experimental evidence for the helical structure of M1. They also suggest that M1 may slightly unwind or have a kink at a Gly in the middle of M1. A model in which M1 from each subunit forms a packing shell around the pore‐forming M2 helices may account for our observations. This work was supported in part by grants from the NIH (DK054354). OBK was supported by NIH grants DK061296 and DK066883. KMB was supported by NIH grant GM60582.