Kinetic interconversion of rat and bovine homologs of the alpha subunit of an amiloride-sensitive Na+ channel by C-terminal truncation of the bovine subunit.
We have recently cloned the alpha subunit of a bovine amiloride-sensitive Na+ channel (alphabENaC). This subunit shares extensive homology with both rat and human alphaENaC subunits but shows marked divergence at the C terminus beginning at amino acid 584 of the 697-residue sequence. When incorporated into planar lipid bilayers, alphabENaC almost exclusively exhibits a main transition to 39 picosiemens (pS) with very rare 13 pS step transitions to one of two subconductance states (26 and 13 pS). In contrast, the alpha subunit of the rat renal homolog of ENaC (alpharENaC) has a main transition step to 13 pS that is almost constituitively open, with a second stepwise transition of 26 to 39 pS. A deletion mutant of alphabENaC, encompassing the entire C-terminal region (R567X), converts the kinetic behavior of alphabENaC to that of alpharENaC, i. e. a transition to 13 pS followed by a second 26 pS transition to 39 pS. Chemical cross-linking of R567X restores the wild-type alphabENaC gating pattern, whereas treatment with the reducing agent dithiothreitol produced only 13 pS transitions. In contrast, an equivalent C-terminal truncation of alpharENaC (R613X) had no effect on the gating pattern of alpharENaC. These results are consistent with the hypothesis that interactions between the C termini of alphabENaC account for the different kinetic behavior of this member of the ENaC family of Na+ channels.
Fuller
C M
CM
Department of Physiology and Biophysics, University of Alabama at Birmingham, Birmingham, Alabama 35294, USA.
Ismailov
I I
II
Berdiev
B K
BK
Shlyonsky
V G
VG
Benos
D J
DJ
D000584:Amiloride / Q000494:pharmacology*; D000595:Amino Acid Sequence; D000818:Animals; D002417:Cattle; D003432:Cross-Linking Reagents; D018076:DNA, Complementary; D006801:Humans; D015640:Ion Channel Gating; D007700:Kinetics; D008051:Lipid Bilayers; D008969:Molecular Sequence Data; D016296:Mutagenesis; D016366:Open Reading Frames; D009994:Osmolar Concentration; D051381:Rats; D017386:Sequence Homology, Amino Acid; D015222:Sodium Channels / Q000737:chemistry* / Q000187:drug effects / Q000235:genetics; D012965:Sodium Chloride; D013439:Sulfhydryl Reagents; D014981:Xenopus
D003432:Cross-Linking Reagents; D018076:DNA, Complementary; D008051:Lipid Bilayers; D015222:Sodium Channels; D013439:Sulfhydryl Reagents; D012965:Sodium Chloride; D000584:Amiloride