Constraint-based, homology model of the extracellular domain of the epithelial Na+ channel α subunit reveals a mechanism of channel activation by proteases.
The epithelial Na(+) channel (ENaC) mediates Na(+) transport across high resistance epithelia. This channel is assembled from three homologous subunits with the majority of the protein's mass found in the extracellular domains. Acid-sensing ion channel 1 (ASIC1) is homologous to ENaC, but a key functional domain is highly divergent. Here we present molecular models of the extracellular region of α ENaC based on a large data set of mutations that attenuate inhibitory peptide binding in combination with comparative modeling based on the resolved structure of ASIC1. The models successfully rationalized the data from the peptide binding screen. We engineered new mutants that had not been tested based on the models and successfully predict sites where mutations affected peptide binding. Thus, we were able to confirm the overall general fold of our structural models. Further analysis suggested that the α subunit-derived inhibitory peptide affects channel gating by constraining motions within two major domains in the extracellular region, the thumb and finger domains.
Department of Medicine, University of Pittsburgh, Pittsburgh, Pennsylvania 15261, USA.
Ossama B
OB
Kashlan
Joshua L
JL
Adelman
Sora
S
Okumura
Brandon M
BM
Blobner
Zachary
Z
Zuzek
Rebecca P
RP
Hughey
Thomas R
TR
Kleyman
Michael
M
Grabe
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D062686:Epithelial Sodium Channel Blockers; D053503:Epithelial Sodium Channels / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D005110:Extracellular Space / Q000378:metabolism*; D045683:Furin / Q000378:metabolism; D015640:Ion Channel Gating; D051379:Mice; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009068:Movement; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D010447:Peptide Hydrolases / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D017386:Sequence Homology, Amino Acid
D062686:Epithelial Sodium Channel Blockers; D053503:Epithelial Sodium Channels; D010447:Peptide Hydrolases; D045683:Furin