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Bases Structurales De La Specificite De Reconnaissance Entre Les Toxines Animales Et Les Canaux Ioniques Cedric Bernard

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Bases Structurales De La Specificite De Reconnaissance Entre Les Toxines Animales Et Les Canaux Ioniques Cedric Bernard Bases structurales de la specificite de reconnaissance entre les toxines animales et les canaux ioniques Cedric Bernard To cite this version: Cedric Bernard. Bases structurales de la specificite de reconnaissance entre les toxines animales et les canaux ioniques. Biochimie [q-bio.BM]. Université de la Méditerranée - Aix-Marseille II, 2002. Français. tel-00010404 HAL Id: tel-00010404 https://tel.archives-ouvertes.fr/tel-00010404 Submitted on 5 Oct 2005 HAL is a multi-disciplinary open access L’archive ouverte pluridisciplinaire HAL, est archive for the deposit and dissemination of sci- destinée au dépôt et à la diffusion de documents entific research documents, whether they are pub- scientifiques de niveau recherche, publiés ou non, lished or not. The documents may come from émanant des établissements d’enseignement et de teaching and research institutions in France or recherche français ou étrangers, des laboratoires abroad, or from public or private research centers. publics ou privés. Remerciements Je tiens tout d’abord à remercier les membres du jury qui ont accepté de juger ce travail. Je remercie christian Cambillau de m’avoir permis d’être intégré au sein de l’A.F.M.B. Une profonde pensée pour ma mère, mais aussi pour ma sœur Karin (avec toute ta petite famille vivant au rythme du zouc antillais), pour Thomas et Geoffroy mes frères enfin retrouvés. Merci à tous de si bien m’entourer même si j’ai des fois l’impression de ne pas le mériter… J’aimerais rendre hommage à la famille Gabella, famille dont je me targue d’appartenir un peu. Vous avez tant fait pour moi que je ne pourrai jamais assez vous remercier. Initialement protecteurs et guides, vous me faites désormais l’honneur de donner votre amitié à l’homme que je suis devenu. Je n’oublierai jamais ces instants précieux… En ce qui concerne mes amis, j’ai tellement peur d’oublier quelqu’un que je me contenterai de leur offrir un exemplaire dédicacé de ma thèse (…et donc, exemplaire unique en quelque sorte…). J’aimerais aussi remercier tous les membres du laboratoire et plus particulièrement mes camarades de RMN : Eric, Olivier, Gilles, Jean- guillaume, Guillaume et Benjamin. Notre entente et votre (mon ?) humour m’ont fait passer quatre années excellentes, aussi bien en terme de travail qu’en terme de détente…. Enfin, il me reste à remercier Hervé Darbon. Exercice difficile puisqu’il faut ainsi dissocier l’homme du directeur de thèse. Force m’est de constater que ces deux faces du personnage sont entremêlées, sans césure, en un mot indissociables. L’attitude très paternelle que tu adoptes vis-à-vis des étudiants de ton groupe, ou plutôt de ton clan RMN fait chaud au cœur et me manquera. Merci de m’avoir formé : ce que je sais aujourd’hui, je te le dois en partie. Merci de m’avoir dirigé et permis d’aller assister à des congrès. Merci pour ces instants de grande détente sur ton bateau. Merci pour tes « tu es à la bourre Cédric », pour tes encouragements, pour tes blagues qui ne me faisaient pas rire. Enfin, merci pour ton amitié : si une chose, une seule devait perdurer, j’espère que ce serait celle la… Et Florence … I - II - Avant propos Les canaux ioniques sont exprimés de façon ubiquitaire et occupent donc une place primordiale dans les mécanismes du vivant. De fait, un grand nombre de pathologies impliquent un dysfonctionnement d’un certain type de canal ionique, et motivent ainsi l’énorme quantité d’études qui visent à mieux comprendre ces canaux. Dans l’optique d‘envisager un traitement efficace de ces maladies, la compréhension des mécanismes de fonctionnement des canaux ioniques constitue un objectif majeur à atteindre. Les toxines animales, initialement étudiées pour obtenir des anti-venins efficaces, apparaissent aujourd’hui comme un outil de choix pour étudier et mieux comprendre le fonctionnement des canaux ioniques. Le travail effectué au cours de cette thèse est sous-tendu par une idée maîtresse, améliorer encore la connaissance et la compréhension de l’interaction d’une toxine et de sa cible, le canal. Ces connaissances ont diverses répercussions : la caractérisation encore plus fine des propriétés des différents types de canaux exprimés mais aussi la création de molécules thérapeutiques à la spécificité contrôlée pour un certain type de canal donné. Ce contrôle de la spécificité, qui doit être total, implique une connaissance parfaite des caractéristiques physico-chimiques qui régissent cette spécificité. Les connaissances en la matière sont maintenant nombreuses, mais les zones d’ombre, qu’il reste à dévoiler, le sont plus encore. La mutagénèse dirigée, la création de molécules chimériques et les tests électrophysiologiques ont été largement utilisés pour mieux cerner les résidus critiques pour l’interaction de la toxine et affiner ainsi le modèle de l’interaction entre une toxine et un canal ionique. Toutefois, l’émergence et l’utilisation désormais courante des techniques de détermination de la structure des protéines ouvre un nouveau champ d’analyse qui se base sur la comparaison des structures et plus précisément permet l’établissement de véritables cartes fonctionnelles de ces toxines envers un type de canal donné. Au travers de son introduction bibliographique, ce manuscrit présente brièvement les canaux ioniques en se focalisant sur les relations structure/fonction de ces protéines membranaires. Puis les toxines animales, dont les canaux ioniques sont les cibles naturelles, sont décrites suivant le motif structural qu’elles adoptent : soit le motif Csαβ ou le motif ICK. Enfin, les résultats obtenus s’articulent autour de la détermination de la structure en solution de toxines animales par résonance magnétique nucléaire bidimensionnelle du proton. - III - - IV - Table des matières INTRODUCTION 1ERE PARTIE : LES CANAUX IONIQUES................................................................ 3 I Les canaux potassium ....................................................................................... 7 1 LES TROIS GRANDES CLASSES DE CANAUX POTASSIUM.......................................... 7 a°) Les canaux potassium activés par le calcium (KCA)................................................................. 7 i Les KCA de grande conductance (canaux BK) .......................................................... 8 ii Les KCA de conductance intermédiaire (canaux IK)................................................. 8 iii Les KCA de faible conductance (canaux SK)............................................................. 8 b°) Les canaux potassium dépendants du potentiel (Kv)................................................................ 9 c°) Les canaux rectifiants entrants (canal KIR)............................................................................. 10 2 ORGANISATION MOLECULAIRE DES CANAUX POTASSIUM .................................... 10 a°) Les segments transmembranaires........................................................................................... 10 b°) Une séquence signature : le filtre de sélectivité ..................................................................... 11 c°) La sensibilité au potentiel....................................................................................................... 12 + 3 LA STRUCTURE TRIDIMENSIONNELLE DU CANAL K BACTERIEN KCSA............... 12 a°) Organisation moléculaire du canal KcsA............................................................................... 13 b°) Un modèle de la conduction d’ions K+.................................................................................. 14 c°) Les réarrangements conformationnels ................................................................................... 15 4 LA STRUCTURE TRIDIMENSIONNELLE DU CANAL MTHK...................................... 16 II Les canaux calcium voltage-dépendants .......................................................... 19 1 LA DIVERSITE DES CANAUX CALCIUM VOLTAGE-DEPENDANTS ........................... 19 a°) Les canaux Ca2+ de type LVA................................................................................................ 19 b°) Les canaux Ca2+ de type HVA ............................................................................................... 20 i Les canaux de type L .............................................................................................. 20 ii Les canaux de type N.............................................................................................. 21 iii Les canaux de type P/Q .......................................................................................... 21 iv Les canaux de type R .............................................................................................. 21 c°) Une nouvelle classification .................................................................................................... 22 2 ORGANISATION MOLECULAIRE DES VSCC........................................................... 23 3 LA PERMEATION SELECTIVE DES IONS CALCIUM .................................................. 25 III Les canaux sodium.......................................................................................... 28 1 CLASSIFICATION DES CANAUX SODIUM................................................................ 28 2 ORGANISATION MOLECULAIRE DES CANAUX SODIUM.......................................... 28 a°) Topologie des canaux sodium................................................................................................ 28 b°) Sélectivité et perméation .......................................................................................................
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