Functional Roles of Clusters of Hydrophobic and Polar Residues in the Epithelial Na+ Channel Knuckle Domain.

ABSTRACT: The extracellular regions of epithelial Na(+) channel subunits are highly ordered structures composed of domains formed by ? helices and ? strands. Deletion of the peripheral knuckle domain of the ? subunit in the ??? trimer results in channel activation, reflecting an increase in channel open probability due to a loss of the inhibitory effect of external Na(+) (Na(+) self-inhibition). In contrast, deletion of either the ? or ? subunit knuckle domain within the ??? trimer dramatically reduces epithelial Na(+) channel function and surface expression, and impairs subunit maturation. We systematically mutated individual ? subunit knuckle domain residues and assessed functional properties of these mutants. Cysteine substitutions at 14 of 28 residues significantly suppressed Na(+) self-inhibition. The side chains of a cluster of these residues are non-polar and are predicted to be directed toward the palm domain, whereas a group of polar residues are predicted to orient their side chains toward the space between the knuckle and finger domains. Among the mutants causing the greatest suppression of Na(+) self-inhibition were ?P521C, ?I529C, and ?S534C. The introduction of Cys residues at homologous sites within either the ? or ? subunit knuckle domain resulted in little or no change in Na(+) self-inhibition. Our results suggest that multiple residues in the ? subunit knuckle domain contribute to the mechanism of Na(+) self-inhibition by interacting with palm and finger domain residues via two separate and chemically distinct motifs.

SUBMITTER: Chen J 

PROVIDER: S-EPMC4599017 | biostudies-literature | 2015 Oct

REPOSITORIES: biostudies-literature