Refining a structural model of ENaC

University Of Pittsburgh At Pittsburgh, Pittsburgh PA

Investigators

Linked publications & trials

Abstract

DESCRIPTION (provided by applicant): The epithelial Na+ channel (ENaC) has a key role in the regulation of extracellular fluid volume and blood pressure. ENaC belongs to a family of ion channels that sense the external environment. ENaC responds to several environmental cues including Na+, Cl-, protons, proteases and shear stress. The ENaC extracellular domains interact with the extracellular environment to sense these cues. Having an accurate model of ENaC structure is therefore critical to understanding ENaC regulation by extracellular factors. Acid sensing ion channel 1 (ASIC1) is the only protein homologous to ENaC whose structure has been reported. The ENaC subunit extracellular domains are largely homologous the corresponding ASIC1 domains, except for the so called finger domain where ENaC subunits have a large insertion. The ENaC finger domain has been implicated in the channel's response to Na+, proteases, and shear stress. The goal of this proposal is to improve upon a structural model of the extracellular domains of the ENaC ? subunit. The proposed studies will investigate the structural interactions that involve a stretch of residues that includes two cysteines in the finger domain insertion. We will then use this information to refine a structural model of the ENaC ? subunit. We previously hypothesized that these two finger domain cysteines formed a disulfide bridged pair, but our preliminary data contradicts this notion. Experiments are proposed to define specific interactions between these cysteines and other specific sites in the channel, and between these cysteines and an inhibitory peptide. We will derive distance constraints from our data and combine them with constraints based on homology to ASIC1 and on other available data to refine a model of the ENaC ? subunit extracellular domains. We will then assess our model by developing novel hypotheses of function to test.