Structure of the 5th transmembrane segment of the Na,K-ATPase alpha subunit: a cysteine-scanning mutagenesis study.
To study the structure of the pathway of cations across the Na, K-ATPase, we applied the substituted cysteine accessibility method to the putative 5th transmembrane segment of the alpha subunit of the Na,K-ATPase of the toad Bufo marinus. Only the most extracellular amino acid position (A(796)) was accessible from the extracellular side in the native Na,K-pump. After treatment with palytoxin, six other positions (Y(778), L(780), S(782), P(785), E(786) and L(791)), distributed along the whole length of the segment, became readily accessible to a small-size methanethiosulfonate compound (2-aminoethyl methanethiosulfonate). The accessible residues are not located on the same side of an alpha-helical model but the pattern of reactivity would rather suggest a beta-sheet structure for the inner half of the putative transmembrane segment. These results demonstrate the contribution of the 5th transmembrane segment to the palytoxin-induced channel and indicate which amino acid positions are exposed to the pore of this channel.
Guennoun
S
S
Institute of Pharmacology and Toxicology, Bugnon 27, CH-1005 Lausanne, Switzerland.
Horisberger
J D
JD
D000178:Acrylamides / Q000494:pharmacology; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D002024:Bufo marinus; D002462:Cell Membrane / Q000201:enzymology; D003064:Cnidarian Venoms / Q000494:pharmacology; D003545:Cysteine; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009865:Oocytes / Q000502:physiology; D017433:Protein Structure, Secondary; D021122:Protein Subunits; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D000254:Sodium-Potassium-Exchanging ATPase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D010566:Virulence Factors, Bordetella / Q000494:pharmacology; D000095702:Polyether Toxins
D000178:Acrylamides; D003064:Cnidarian Venoms; D003545:Cysteine; D021122:Protein Subunits; D011994:Recombinant Proteins; D000254:Sodium-Potassium-Exchanging ATPase; D010566:Virulence Factors, Bordetella; C010272:palytoxin; D000095702:Polyether Toxins