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Article: Helical secondary structure of the external S3-S4 linker of pacemaker (HCN) channels revealed by site-dependent perturbations of activation phenotype

TitleHelical secondary structure of the external S3-S4 linker of pacemaker (HCN) channels revealed by site-dependent perturbations of activation phenotype
Authors
KeywordsChemicals And Cas Registry Numbers
Issue Date2003
PublisherAmerican Society for Biochemistry and Molecular Biology, Inc. The Journal's web site is located at http://www.jbc.org/
Citation
Journal Of Biological Chemistry, 2003, v. 278 n. 25, p. 22290-22297 How to Cite?
AbstractIf, encoded by the hyperpolarization-activated cyclic nucleotide-modulated channel family (HCN1-4), contributes significantly to neuronal and cardiac pacing. Recently, we reported that the S3-S4 residue Glu-235 of HCN1 influences activation by acting as a surface charge. However, it is uncertain whether other residues of the external S3-S4 linker are also involved in gating. Furthermore, the secondary conformation of the linker is not known. Here we probed the structural and functional role of the HCN1 S3-S4 linker by introducing systematic mutations into the entire linker (defined as 229-237) and studying their effects. We found that the mutations K230A (-62.2 ± 3.4 mV versus -72.2 ± 1.7 mV of wild type (WT)), G231A (-64.4 ± 1.3 mV), M232A (V1/2 = -63.1 ± 1.1 mV), and E235G (-65.4 ± 1.5 mV) produced depolarizing activation shifts. Although E229A and M232A decelerated gating kinetics (<13- and 3-fold, respectively), K230A and G231A accelerated both activation and deactivation (<∼2-3-fold). D233A, S234A, V236A, and Y237A channels exhibited WT properties (p > 0.05). Shortening the linker (EVY235-237ΔΔΔ) caused depolarizing activation shift and slowed kinetics that could not be explained by removing the charge at position 235 alone. Secondary structural predictions by the modeling algorithms SSpro2 and PROF, along with refinements by our experimental data, suggest that part of the S3-S4 linker conforms a helical structure with the functionally important residues Met-232, Glu-235, and Gly-231 (|ΔΔG|>1 kcal/mol) clustered on one side.
Persistent Identifierhttp://hdl.handle.net/10722/91483
ISSN
2015 Impact Factor: 4.258
2015 SCImago Journal Rankings: 3.151
ISI Accession Number ID
References

 

DC FieldValueLanguage
dc.contributor.authorLesso, Hen_HK
dc.contributor.authorLi, RAen_HK
dc.date.accessioned2010-09-17T10:20:08Z-
dc.date.available2010-09-17T10:20:08Z-
dc.date.issued2003en_HK
dc.identifier.citationJournal Of Biological Chemistry, 2003, v. 278 n. 25, p. 22290-22297en_HK
dc.identifier.issn0021-9258en_HK
dc.identifier.urihttp://hdl.handle.net/10722/91483-
dc.description.abstractIf, encoded by the hyperpolarization-activated cyclic nucleotide-modulated channel family (HCN1-4), contributes significantly to neuronal and cardiac pacing. Recently, we reported that the S3-S4 residue Glu-235 of HCN1 influences activation by acting as a surface charge. However, it is uncertain whether other residues of the external S3-S4 linker are also involved in gating. Furthermore, the secondary conformation of the linker is not known. Here we probed the structural and functional role of the HCN1 S3-S4 linker by introducing systematic mutations into the entire linker (defined as 229-237) and studying their effects. We found that the mutations K230A (-62.2 ± 3.4 mV versus -72.2 ± 1.7 mV of wild type (WT)), G231A (-64.4 ± 1.3 mV), M232A (V1/2 = -63.1 ± 1.1 mV), and E235G (-65.4 ± 1.5 mV) produced depolarizing activation shifts. Although E229A and M232A decelerated gating kinetics (<13- and 3-fold, respectively), K230A and G231A accelerated both activation and deactivation (<∼2-3-fold). D233A, S234A, V236A, and Y237A channels exhibited WT properties (p > 0.05). Shortening the linker (EVY235-237ΔΔΔ) caused depolarizing activation shift and slowed kinetics that could not be explained by removing the charge at position 235 alone. Secondary structural predictions by the modeling algorithms SSpro2 and PROF, along with refinements by our experimental data, suggest that part of the S3-S4 linker conforms a helical structure with the functionally important residues Met-232, Glu-235, and Gly-231 (|ΔΔG|>1 kcal/mol) clustered on one side.en_HK
dc.languageengen_HK
dc.publisherAmerican Society for Biochemistry and Molecular Biology, Inc. The Journal's web site is located at http://www.jbc.org/en_HK
dc.relation.ispartofJournal of Biological Chemistryen_HK
dc.subjectChemicals And Cas Registry Numbersen_HK
dc.subject.meshAlgorithmsen_HK
dc.subject.meshAmino Acid Sequenceen_HK
dc.subject.meshAmino Acid Substitutionen_HK
dc.subject.meshAnimalsen_HK
dc.subject.meshBinding Sitesen_HK
dc.subject.meshCloning, Molecularen_HK
dc.subject.meshCyclic Nucleotide-Gated Cation Channelsen_HK
dc.subject.meshIon Channels - chemistry - physiologyen_HK
dc.subject.meshMembrane Potentials - physiologyen_HK
dc.subject.meshMiceen_HK
dc.subject.meshModels, Molecularen_HK
dc.subject.meshMutagenesis, Site-Directeden_HK
dc.subject.meshNerve Tissue Proteinsen_HK
dc.subject.meshPatch-Clamp Techniquesen_HK
dc.subject.meshPolymerase Chain Reactionen_HK
dc.subject.meshPotassium Channelsen_HK
dc.subject.meshProtein Structure, Secondaryen_HK
dc.subject.meshRecombinant Proteins - chemistryen_HK
dc.subject.meshSequence Deletionen_HK
dc.titleHelical secondary structure of the external S3-S4 linker of pacemaker (HCN) channels revealed by site-dependent perturbations of activation phenotypeen_HK
dc.typeArticleen_HK
dc.identifier.emailLi, RA:ronaldli@hkucc.hku.hken_HK
dc.identifier.authorityLi, RA=rp01352en_HK
dc.description.naturelink_to_subscribed_fulltext-
dc.identifier.doi10.1074/jbc.M302466200en_HK
dc.identifier.pmid12668666-
dc.identifier.scopuseid_2-s2.0-0038492590en_HK
dc.relation.referenceshttp://www.scopus.com/mlt/select.url?eid=2-s2.0-0038492590&selection=ref&src=s&origin=recordpageen_HK
dc.identifier.volume278en_HK
dc.identifier.issue25en_HK
dc.identifier.spage22290en_HK
dc.identifier.epage22297en_HK
dc.identifier.isiWOS:000183503900017-
dc.publisher.placeUnited Statesen_HK
dc.identifier.scopusauthoridLesso, H=6507506626en_HK
dc.identifier.scopusauthoridLi, RA=7404724466en_HK

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