Home | Help | Feedback | Subscriptions | Archive | Search | Table of Contents

This Article Full Text Full Text (PDF, 170K) PPT slides of all figures Alert me when this article is cited Citation Map Services Email this article Similar articles in this journal Similar articles in PubMed Alert me to new content in the JGP Download to citation manager Citing Articles Citing Articles via CrossRef Citing Articles via Google Scholar Google Scholar Articles by Heginbotham, L. Articles by Miller, C. Search for Related Content PubMed PubMed Citation Articles by Heginbotham, L. Articles by Miller, C. Pubmed/NCBI databases Compound via MeSH Substance via MeSH Social Bookmarking                            What's this?

Original Article

Functional Asymmetries and Sidedness of Proton Activation

Lise Heginbotham^a, Meredith LeMasurier^a, Ludmilla Kolmakova-Partensky^a, and Christopher Miller^a

^a From the Department of Biochemistry, Howard Hughes Medical Institute, Brandeis University, Waltham, Massachusetts 02254-9110
Department of Biochemistry, Brandeis University, 415 South Street, HHMI, Waltham, MA 02254-9110.Fax: 781-736-2365;

cmiller{at}brandeis.edu

Basic electrophysiological properties of the KcsA K⁺ channel were examined in planar lipid bilayer membranes. The channel displays open-state rectification and weakly voltage-dependent gating. Tetraethylammonium blocking affinity depends on the side of the bilayer to which the blocker is added. Addition of Na⁺ to the trans chamber causes block of open-channel current, while addition to the cis side has no effect. Most striking is the activation of KcsA by protons; channel activity is observed only when the trans bilayer chamber is at low pH. To ascertain which side of the channel faces which chamber, residues with structurally known locations were mapped to defined sides of the bilayer. Mutation of Y82, an external residue, results in changes in tetraethylammonium affinity exclusively from the cis side. Channels with cysteine residues substituted at externally exposed Y82 or internally exposed Q119 are functionally modified by methanethiosulfonate reagents from the cis or trans chambers, respectively. Block by charybdotoxin, known to bind to the channel's external mouth, is observed only when the toxin is added to the cis side of channels mutated to be toxin sensitive. These results demonstrate unambiguously that the protonation sites linked to gating are on the intracellular portion of the KcsA protein.

Key Words: potassium channel • permeation • gating • block

CiteULike    Complore    Connotea    Del.icio.us    Digg    Facebook    Reddit    Technorati    Twitter    What's this?

Home | Help | Feedback | Subscriptions | Archive | Search | Table of Contents