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1. NMR studies of the interaction of calmodulin with IQ motif peptides

Author

Walter J. Chazin, Benjamin Chagot, Steven M. Damo, Michael D. Feldkamp, Center for Structural Biology, Vanderbilt University [Nashville], Ingénierie Moléculaire et Physiopathologie Articulaire ( IMoPA ), Université de Lorraine ( UL ) -Centre National de la Recherche Scientifique ( CNRS ), Ingénierie Moléculaire et Physiopathologie Articulaire (IMoPA), Université de Lorraine (UL)-Centre National de la Recherche Scientifique (CNRS), and Centre National de la Recherche Scientifique (CNRS)-Université de Lorraine (UL)

Subjects
MESH: Signal Transduction, MESH : NAV1.5 Voltage-Gated Sodium Channel, Amino Acid Motifs, Intracellular Space, MESH : Intracellular Space, Peptide, Plasma protein binding, EF-hand, Na 15, 15 N-1 H HSQC, IQ motif, NAV1.5 Voltage-Gated Sodium Channel, MESH: Amino Acid Motifs, MESH: Apoproteins, Calcium-binding protein, MESH: Nuclear Magnetic Resonance, Biomolecular, MESH: Peptide Fragments, Calcium signaling, chemistry.chemical_classification, 0303 health sciences, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], biology, EF hand, 030302 biochemistry & molecular biology, MESH : Apoproteins, MESH : Protein Binding, Nuclear magnetic resonance spectroscopy, MESH: Calmodulin, 3. Good health, [SDV.BBM.BP]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biophysics, Biochemistry, MESH: Calcium, MESH: Intracellular Space, Protein Binding, Signal Transduction, MESH : Calmodulin, [ SDV.BBM.BP ] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biophysics, MESH : Amino Acid Motifs, animal structures, Calmodulin, MESH : Nuclear Magnetic Resonance, Biomolecular, calcium signaling, Article, 03 medical and health sciences, NMR spectroscopy, MESH: Protein Binding, Humans, MESH : Calcium, Nuclear Magnetic Resonance, Biomolecular, Ion channel, 030304 developmental biology, MESH : Signal Transduction, MESH: Humans, protein complex, MESH : Peptide Fragments, MESH : Humans, MESH: NAV1.5 Voltage-Gated Sodium Channel, calcium binding protein, Peptide Fragments, co-expression, chemistry, biology.protein, Calcium, Apoproteins, [ SDV.BBM.BS ] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM]
Abstract

International audience; Calmodulin (CaM) is a ubiquitous EF-hand calcium sensor protein that transduces calcium signals in a wide range of signaling pathways. Structural analysis of complexes with peptides has provided valuable insights into the remarkable variety in the way in which CaM interacts with and activates its targets. Among these various targets, CaM has been shown to be an essential component of a calcium-sensing regulatory apparatus for a number of voltage-gated ion channels. NMR spectroscopy has proven to be a powerful tool for the structural characterization of CaM-peptide complexes, in particular for the study of IQ motifs, which bind CaM at the basal level of calcium in cells and thereby serve to localize CaM to its sites of action. We describe here methods for the robust expression and purification of CaM isotopically enriched for NMR analysis, as well as for the complex of CaM with a peptide derived from the IQ motif sequence of the human cardiac sodium channel Na(V)1.5. We also describe methods for NMR analysis of titrations of CaM with IQ motif peptides to determine the stoichiometry of the complex and to identify the residues at the binding interface.

Published
2013
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2. Parental electrocardiographic screening identifies a high degree of inheritance for congenital and childhood nonimmune isolated atrioventricular block

Author

Hervé Le Marec, Caroline Bonnet, Jean-René Lusson, Raphaël P. Martins, Jean-Jacques Schott, Francis Rouault, Alban-Elouen Baruteau, Jean-Benoit Thambo, Christian Dina, Elisabeth Villain, François Godart, Philippe Mabo, Vincent Probst, Alain Fraisse, François Marçon, Albin Behaghel, Véronique Gournay, Sophie Guillaumont, Yves Dulac, Jean-Marc Schleich, Stéphanie Chatel, Swanny Fouchard, Alain Chantepie, Béatrice Delasalle, Jean-Claude Daubert, unité de recherche de l'institut du thorax UMR1087 UMR6291 ( ITX ), Université de Nantes ( UN ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Centre National de la Recherche Scientifique ( CNRS ), Department de Chirurgie cardiaque des cardiopathies congénitales, Centre chirurgical Marie Lannelongue, Service de cardiologie et maladies vasculaires, Université de Rennes 1 ( UR1 ), Université de Rennes ( UNIV-RENNES ) -Université de Rennes ( UNIV-RENNES ) -Hôpital Pontchaillou-CHU Pontchaillou [Rennes], CIC-IT Rennes, Hôpital Pontchaillou-Institut National de la Santé et de la Recherche Médicale ( INSERM ), Laboratoire Traitement du Signal et de l'Image ( LTSI ), Université de Rennes ( UNIV-RENNES ) -Université de Rennes ( UNIV-RENNES ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ), Laboratoire de génie des procédés - environnement - agroalimentaire ( GEPEA ), Mines Nantes ( Mines Nantes ) -Université de Nantes ( UN ) -Ecole Nationale Vétérinaire, Agroalimentaire et de l'alimentation Nantes-Atlantique ( ONIRIS ) -Centre National de la Recherche Scientifique ( CNRS ), Service de cardiologie pédiatrique [CHU Necker], Assistance publique - Hôpitaux de Paris (AP-HP)-CHU Necker - Enfants Malades [AP-HP], Service cardiologie pédiatrique [Bordeaux], CHU Bordeaux [Bordeaux], Service de Cardiologie Infantile [CHRU Nancy], Centre Hospitalier Régional Universitaire de Nancy ( CHRU Nancy ), Service de cardiologie pédiatrique [Nantes], Centre hospitalier universitaire de Nantes ( CHU Nantes ), Cabinet de Cardiologie pédiatrique, Service de cardiologie pédiatrique [Tours], CHU Tours, Service de cardiologie Pédiatrique [Montpellier], Centre Hospitalier Régional Universitaire [Montpellier] ( CHRU Montpellier ), Service de cardiologie Pédiatrique [Lille], Centre Hospitalier Régional Universitaire [Lille] ( CHRU Lille ), Service de cardiologie pédiatrique et congénitale adulte [CHU de Dijon], Centre Hospitalier Universitaire de Dijon - Hôpital François Mitterrand ( CHU Dijon ), Service de cardiologie Pédiatrique [Marseille], Hôpital de la Timone [CHU - APHM] ( TIMONE ), Service de cardiologie [Clermont-Ferrand], CHU Clermont-Ferrand, Service pédiatrie-cardiologie, CHU Toulouse [Toulouse]-Hôpital des Enfants, unité de recherche de l'institut du thorax UMR1087 UMR6291 (ITX), Université de Nantes - UFR de Médecine et des Techniques Médicales (UFR MEDECINE), Université de Nantes (UN)-Université de Nantes (UN)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Service de cardiologie et maladies vasculaires [Rennes] = Cardiac, Thoracic, and Vascular Surgery [Rennes], CHU Pontchaillou [Rennes], Hôpital Pontchaillou-Institut National de la Santé et de la Recherche Médicale (INSERM), Laboratoire Traitement du Signal et de l'Image (LTSI), Université de Rennes 1 (UR1), Université de Rennes (UNIV-RENNES)-Université de Rennes (UNIV-RENNES)-Institut National de la Santé et de la Recherche Médicale (INSERM), Laboratoire de génie des procédés - environnement - agroalimentaire (GEPEA), Mines Nantes (Mines Nantes)-Université de Nantes - UFR des Sciences et des Techniques (UN UFR ST), Université de Nantes (UN)-Université de Nantes (UN)-Ecole Nationale Vétérinaire, Agroalimentaire et de l'alimentation Nantes-Atlantique (ONIRIS)-Centre National de la Recherche Scientifique (CNRS), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-CHU Necker - Enfants Malades [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Centre Hospitalier Régional Universitaire de Nancy (CHRU Nancy), Centre hospitalier universitaire de Nantes (CHU Nantes), Centre Hospitalier Régional Universitaire de Tours (CHRU Tours), Centre Hospitalier Régional Universitaire [Montpellier] (CHRU Montpellier), Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille), Centre Hospitalier Universitaire de Dijon - Hôpital François Mitterrand (CHU Dijon), Hôpital de la Timone [CHU - APHM] (TIMONE), Service de Cardiologie Maladies Vasculaires [CHU Clermont-Ferrand], CHU Gabriel Montpied [Clermont-Ferrand], CHU Clermont-Ferrand-CHU Clermont-Ferrand, CHU Toulouse [Toulouse], Unité de recherche de l'institut du thorax (ITX-lab), Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Université de Nantes - UFR de Médecine et des Techniques Médicales (UFR MEDECINE), Université de Nantes (UN)-Université de Nantes (UN), Centre Chirurgical Marie Lannelongue (CCML), Université de Rennes (UR)-Institut National de la Santé et de la Recherche Médicale (INSERM), Université de Nantes (UN)-Université de Nantes (UN)-École nationale vétérinaire, agroalimentaire et de l'alimentation Nantes-Atlantique (ONIRIS)-Centre National de la Recherche Scientifique (CNRS), Service Cardiologie pédiatrique [CHU Toulouse], Pôle Enfants [CHU Toulouse], Centre Hospitalier Universitaire de Toulouse (CHU Toulouse)-Centre Hospitalier Universitaire de Toulouse (CHU Toulouse), Le Corre, Morgane, and Centre Hospitalier Régional Universitaire de Tours (CHRU TOURS)

Subjects
Male, Parents, MESH : Retrospective Studies, Heart malformation, MESH : NAV1.5 Voltage-Gated Sodium Channel, MESH : Prevalence, MESH : Aged, Electrocardiography, 0302 clinical medicine, MESH: Pregnancy, MESH : Child, Pregnancy, Prenatal Diagnosis, MESH: Child, Medicine, Sinus rhythm, genetics, Child, 0303 health sciences, MESH: Middle Aged, MESH: Genetic Testing, MESH : Infant, MESH: Infant, 3. Good health, MESH : Phenotype, MESH: Young Adult, MESH : Electrocardiography, Child, Preschool, Cardiology and Cardiovascular Medicine, medicine.medical_specialty, conduction, MESH : Young Adult, Prenatal diagnosis, MESH: Phenotype, Sudden death, 03 medical and health sciences, [SDV.MHEP.CSC]Life Sciences [q-bio]/Human health and pathology/Cardiology and cardiovascular system, Physiology (medical), MESH : Adolescent, MESH : Genetic Testing, Humans, MESH : Middle Aged, Genetic Testing, MESH: Prenatal Diagnosis, MESH : Prenatal Diagnosis, MESH: Prevalence, Aged, Retrospective Studies, [SDV.IB] Life Sciences [q-bio]/Bioengineering, MESH: Adolescent, MESH: Parents, MESH: Humans, MESH: Child, Preschool, MESH : Humans, Infant, MESH: Adult, MESH: Retrospective Studies, medicine.disease, MESH : Pregnancy, MESH : Genetic Predisposition to Disease, Atrioventricular block, MESH: Female, Pediatrics, ECG screening, 030204 cardiovascular system & hematology, MESH : Child, Preschool, NAV1.5 Voltage-Gated Sodium Channel, atrioventricular block, Prevalence, Mass Screening, MESH : Female, [ SDV.IB ] Life Sciences [q-bio]/Bioengineering, Family history, MESH : Parents, MESH: Aged, MESH: Infant, Newborn, MESH: Genetic Predisposition to Disease, Middle Aged, MESH : Adult, [ SDV.MHEP.CSC ] Life Sciences [q-bio]/Human health and pathology/Cardiology and cardiovascular system, [SDV.MHEP.CSC] Life Sciences [q-bio]/Human health and pathology/Cardiology and cardiovascular system, MESH : Atrioventricular Block, Phenotype, Female, [SDV.IB]Life Sciences [q-bio]/Bioengineering, medicine.symptom, Adult, Adolescent, pediatrics, MESH : Male, MESH : Infant, Newborn, Asymptomatic, Young Adult, MESH: Atrioventricular Block, Genetic Predisposition to Disease, MESH: Mass Screening, PR interval, 030304 developmental biology, MESH : Mass Screening, business.industry, Infant, Newborn, MESH: NAV1.5 Voltage-Gated Sodium Channel, MESH: Male, MESH: Electrocardiography, business
Abstract

Background— The origin of congenital or childhood nonimmune isolated atrioventricular (AV) block remains unknown. We hypothesized that this conduction abnormality in the young may be a heritable disease. Methods and Results— A multicenter retrospective study (13 French referral centers, from 1980–2009) included 141 children with AV block diagnosed in utero, at birth, or before 15 years of age without structural heart abnormalities and without maternal antibodies. Parents and matched control subjects were investigated for family history and for ECG screening. In parents, a family history of sudden death or progressive cardiac conduction defect was found in 1.4% and 11.1%, respectively. Screening ECGs from 130 parents (mean age 42.0±6.8 years, 57 couples) were compared with those of 130 matched healthy control subjects. All parents were asymptomatic and in sinus rhythm, except for 1 with undetected complete AV block. Conduction abnormalities were more frequent in parents than in control subjects, found in 50.8% versus 4.6%, respectively ( P P P P SCN5A mutation screening identified 2 mutations in 2 patients among 97 children. Conclusions— ECG screening in parents of children affected by idiopathic AV block revealed a high prevalence of conduction abnormalities. These results support the hypothesis of an inheritable trait in congenital and childhood nonimmune isolated AV block.

Published
2012
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