http://www.cnr.it/ontology/cnr/individuo/prodotto/ID209008

Dissecting a regulatory calcium-binding site of CLC-K kidney chloride channels (Articolo in rivista)

Type

Prodotto della ricerca (Classe)
Articolo in rivista (Classe)

Label

Dissecting a regulatory calcium-binding site of CLC-K kidney chloride channels (Articolo in rivista) (literal)

Anno

2012-01-01T00:00:00+01:00 (literal)

Http://www.cnr.it/ontology/cnr/pubblicazioni.owl#doi

10.1085/jgp.201210878 (literal)

Alternative label

Gradogna A; Fenollar-Ferrer C; Forrest LR, Pusch M (2012)
Dissecting a regulatory calcium-binding site of CLC-K kidney chloride channels
in The Journal of general physiology; Rockefeller University Press, New York (Stati Uniti d'America)
(literal)

Http://www.cnr.it/ontology/cnr/pubblicazioni.owl#autori

Gradogna A; Fenollar-Ferrer C; Forrest LR, Pusch M (literal)

Pagina inizio

681 (literal)

Pagina fine

695 (literal)

Http://www.cnr.it/ontology/cnr/pubblicazioni.owl#url

http://jgp.rupress.org/content/140/6/681 (literal)

Http://www.cnr.it/ontology/cnr/pubblicazioni.owl#numeroVolume

140 (literal)

Rivista

?The ?Journal of general physiology (Rivista)

Http://www.cnr.it/ontology/cnr/pubblicazioni.owl#numeroFascicolo

6 (literal)

Note

ISI Web of Science (WOS) (literal)

Http://www.cnr.it/ontology/cnr/pubblicazioni.owl#affiliazioni

Istituto di Biofisica, Consiglio Nazionale delle Ricerche, 16149 Genoa, Italy; Computational Structural Biology Group, Max Planck Institute of Biophysics, 60438 Frankfurt am Main, Germany (literal)

Titolo

Dissecting a regulatory calcium-binding site of CLC-K kidney chloride channels (literal)

Abstract

The kidney and inner ear CLC-K chloride channels, which are involved in salt absorption and endolymph production, are regulated by extracellular Ca2+ in the millimolar concentration range. Recently, Gradogna et al. (2010. J. Gen. Physiol. http://dx.doi.org/10.1085/jgp.201010455) identified a pair of acidic residues (E261 and D278) located in the loop between helices I and J as forming a putative intersubunit Ca2+-binding site in hClC-Ka. In this study, we sought to explore the properties of the binding site in more detail. First, we verified that the site is conserved in hClC-Kb and rClC-K1. In addition, we could confer Ca2+ sensitivity to the Torpedo marmorata ClC-0 channel by exchanging its I-J loop with that from ClC-Ka, demonstrating a direct role of the loop in Ca2+ binding. Based on a structure of a bacterial CLC and a new sequence alignment, we built homology models of ClC-Ka. The models suggested additional amino acids involved in Ca2+ binding. Testing mutants of these residues, we could restrict the range of plausible models and positively identify two more residues (E259 and E281) involved in Ca2+ coordination. To investigate cation specificity, we applied extracellular Zn2+, Mg2+, Ba2+, Sr2+, and Mn2+. Zn2+ blocks ClC-Ka as well as its Ca2+-insensitive mutant, suggesting that Zn2+ binds to a different site. Mg2+ does not activate CLC-Ks, but the channels are activated by Ba2+, Sr2+, and Mn2+ with a rank order of potency of Ca2+ > Ba2+ > Sr2+ = Mn2+ for the human CLC-Ks. Dose-response analysis indicates that the less potent Ba2+ has a lower affinity rather than a lower efficacy. Interestingly, rClC-K1 shows an altered rank order (Ca2+ > Sr2+ >> Ba2+), but homology models suggest that residues outside the I-J loop are responsible for this difference. Our detailed characterization of the regulatory Ca2+-binding site provides a solid basis for the understanding of the physiological modulation of CLC-K channel function in the kidney and inner ear. (literal)

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