S3b amino acid residues do not shuttle across the bilayer in voltage-dependent Shaker K+ channels
AUTOR(ES)
Gonzalez, Carlos
FONTE
National Academy of Sciences
RESUMO
In voltage-dependent channels, positive charges contained within the S4 domain are the voltage-sensing elements. The “voltage-sensor paddle” gating mechanism proposed for the KvAP K+ channel has been the subject of intense discussion regarding its general applicability to the family of voltage-gated channels. In this model, the voltage sensor composed of the S3b and the S4 segment shuttles across the lipid bilayer during channel activation. Guided by this mechanism, we assessed here the accessibility of residues in the S3 segment of the Shaker K+ channel by using cysteine-scanning mutagenesis. Mutants expressed robust K+ currents in Xenopus oocytes and reacted with methanethiosulfonate ethyltrimethylammonium in both closed and open conformations of the channel. Because Shaker has a long S3–S4 linker segment, we generated a deletion mutant with only three residues to emulate the KvAP structure. In this short linker mutant, all of the tested residues in the S3b were accessible to methanethiosulfonate ethyltrimethylammonium in both closed and open conformations. Because the S3b moves together with the S4 domain in the paddle model, we tested the effects of deleting two negative charges or adding a positive charge to this region of the channel. We found that altering the S3b net charge does not modify the total gating charge involved in channel activation. We conclude that the S3b segment is always exposed to the external milieu of the Shaker K+ channel. Our results are incompatible with any model involving a large membrane displacement of segment S3b.
ACESSO AO ARTIGO
http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=554844Documentos Relacionados
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