Characterization Of The C-terminal Half Of Human Juvenile Myoclonic Epilepsy Protein Efhc1: Dimer Formation Blocks Ca2+ And Mg2+ Binding To Its Functional Ef-hand.

dc.creatorMurai, Marcelo J
dc.creatorSassonia, Rogério C
dc.creatorZamboni, André H
dc.creatorConte, Fábio F
dc.creatorMartins-de-Souza, Daniel
dc.creatorAparicio, Ricardo
dc.creatorde Oliveira, Marcelo G
dc.creatorLopes-Cendes, Iscia
dc.date2008-Sep
dc.date2015-11-27T13:12:59Z
dc.date2015-11-27T13:12:59Z
dc.date.accessioned2018-03-29T01:07:06Z
dc.date.available2018-03-29T01:07:06Z
dc.identifierArchives Of Biochemistry And Biophysics. v. 477, n. 1, p. 131-8, 2008-Sep.
dc.identifier1096-0384
dc.identifier10.1016/j.abb.2008.06.008
dc.identifierhttp://www.ncbi.nlm.nih.gov/pubmed/18593566
dc.identifierhttp://repositorio.unicamp.br/jspui/handle/REPOSIP/197874
dc.identifier18593566
dc.identifier.urihttp://repositorioslatinoamericanos.uchile.cl/handle/2250/1298107
dc.descriptionHuman EFHC1 is a member of the EF-hand superfamily of Ca(2+)-binding proteins with three DM10 domains of unclear function. Point mutations in the EFHC1 gene are related to juvenile myoclonic epilepsy, a fairly common idiopathic generalized epilepsy. Here, we report the first structural and thermodynamic analyses of the EFHC1C-terminus (residues 403-640; named EFHC1C), comprising the last DM10 domain and the EF-hand motif. Circular dichroism spectroscopy revealed that the secondary structure of EFHC1C is composed by 34% of alpha-helices and 17% of beta-strands. Size exclusion chromatography and mass spectrometry showed that under oxidizing condition EFHC1C dimerizes through the formation of disulfide bond. Tandem mass spectrometry (MS/MS) analysis of peptides generated by trypsin digestion suggests that the Cys575 is involved in intermolecular S-S bond. In addition, DTNB assay showed that each reduced EFHC1C molecule has one accessible free thiol. Isothermal titration calorimetry (ITC) showed that while the interaction between Ca(2+) and EFHC1C is enthalpically driven (DeltaH=-58.6 to -67 kJ/mol and TDeltaS=-22.5 to -31 kJ/mol) the interaction between Mg(2+) and EFHC1C involves an entropic gain, and is approximately 5 times less enthalpically favorable (DeltaH=-11.7 to -14 kJ/mol and TDeltaS=21.9 to 19 kJ/mol) than for Ca(2+) binding. It was also found that under reducing condition Ca(2+) or Mg(2+) ions bind to EFHC1C in a 1/1 molar ratio, while under oxidizing condition this ratio is reduced, showing that EFHC1C dimerization blocks Ca(2+) and Mg(2+) binding.
dc.description477
dc.description131-8
dc.languageeng
dc.relationArchives Of Biochemistry And Biophysics
dc.relationArch. Biochem. Biophys.
dc.rightsfechado
dc.rights
dc.sourcePubMed
dc.subjectAmino Acid Sequence
dc.subjectBase Sequence
dc.subjectBinding Sites
dc.subjectBlotting, Western
dc.subjectCalcium
dc.subjectCalcium-binding Proteins
dc.subjectChromatography, Gel
dc.subjectDna Primers
dc.subjectDimerization
dc.subjectHumans
dc.subjectMagnesium
dc.subjectMass Spectrometry
dc.subjectMolecular Sequence Data
dc.subjectMyoclonic Epilepsy, Juvenile
dc.subjectProtein Binding
dc.subjectProtein Structure, Secondary
dc.titleCharacterization Of The C-terminal Half Of Human Juvenile Myoclonic Epilepsy Protein Efhc1: Dimer Formation Blocks Ca2+ And Mg2+ Binding To Its Functional Ef-hand.
dc.typeArtículos de revistas


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