Structure-function studies of the Kir6.2 channel and of its coupling with natural and artificial partners

157 Pages·2017·8.4 MB·English

by  Maria Antonietta Principalli

Structure-function studies of the Kir6.2 channel and of
its coupling with natural and artificial partners
Maria Antonietta Principalli
To cite this version:
MariaAntoniettaPrincipalli. Structure-functionstudiesoftheKir6.2channelandofitscouplingwith
natural and artificial partners. Molecular biology. Université Grenoble Alpes, 2015. English. ￿NNT:
2015GREAV014￿. ￿tel-01374865￿
HAL Id: tel-01374865
https://theses.hal.science/tel-01374865
Submitted on 2 Oct 2016
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THÈSE
Pour obtenir le grade de
DOCTEUR DE L’UNIVERSITÉ GRENOBLE ALPES
Spécialité : Biologie Structurale et Nanobiologie
Arrêté ministériel : 7 août 2006

Présentée par
Maria Antonietta PRINCIPALLI

Thèse dirigée par Dr. Michel VIVAUDOU et
codirigée par Jean REVILLOUD

préparée au sein de l’Institut de Biologie Structurale
dans l'École Doctorale Chimie et Sciences du Vivant

Étude structure-fonction du canal
Kir6.2 et de son couplage avec des
partenaires naturels et artificiels

Thèse soutenue publiquement le 9 Octobre 2015
devant le jury composé de :

Dr. Pierre-Jean CORRINGER
Président
Prof. Anna MORONI
Rapporteur
Prof. Bruno ALLARD
Rapporteur
Dr. Patrice CATTY
Membre
Dr Michel VIVAUDOU
Directeur de thèse
Jean REVILLOUD
Co-directeur de thèse

Université de Grenoble
École Doctorale Chimie et Science du Vivant
Thèse pour obtenir le grade de
DOCTEUR ès SCIENCES DE L’UNIVERSITÉ DE
GRENOBLE
Specialité: Biologie Structurale et Nanobiologie

Presented and defended in public by
Maria Antonietta PRINCIPALLI
On October 9th 2015

Structure-function studies of the Kir6.2 channel and of
its coupling with natural and artificial partners

Thesis directors:
Dr. Michel VIVAUDOU
Jean REVILLOUD
Jury:
President: Dr. Patrice CATTY
Reviewer: Prof. Anna MORONI
Reviewer: Prof. Bruno ALLARD
Examiner: Dr. Pierre-Jean CORRINGER

Acknowledgements
I would like to thank in the first place Prof. Anna Moroni and Prof. Bruno Allard for
dedicating their time to the evaluation of this work. I would like to express my gratitude as
well to Dr. Patrice Catty, Dr. Pierre-Jean Corringer and Dr. Hugues Nury for being part of
my annual evaluation committee, for their precious advices and their optimistic vision of
problematic projects.
A special thanks to Michel Vivaudou, my thesis director, for his advices and his sense of
humour and for giving me the chance to experience an adventure like no other, scientifically
and personally. Thanks to Jean Revilloud, my supervisor in the lab, for his patience, for
saving my life during my first (and last) day on a pair of skies, and for teaching me the art of
patch-clamping. Many thanks to Christophe Moreau, the most patient person I have ever
known, for encouraging me to believe in my theories and for helping me exploring my ideas
even though they looked crazy or useless to others. Special thanks to Kasia, my personal
English teacher, who helped me in moving the first steps in the lab. Big thanks to Catalina,
whose friendship and Latin spirit helped me reach the end of this adventure. Thanks to Laura
for being an intelligent and fun co-worker, and to Karla for her calm in panic situations.
A particular thanks to Yann, Sonja, Ola, Hubert, Lionel, Céline, Michel T. and Isabelle for
their priceless advices and their help.
Un grazie tutto particolare alla banda degli italiani a Grenoble: il Mollica, ‘mamma tuttofare’,
DdS e mamma Rocha (italiana adottiva), per avermi salvato dai guai in più di un’occasione.
Al piccolo Gabriel per avermi insegnato che non ci si veste mai per bene quando si deve fare
da baby-sitter, a Filippo per il suo accento toscano e le sue email memorabili, a Sciaken per
saper trasmettere la sua calma e il suo approccio positivo alla vita. Grazie a tutti voi ragazzi!
Un grazie speciale a Mizar, con cui ho intrapreso questa e molte altre avventure, amica sincera
e cuoca personale, all’occorrenza pronta a darmi una strigliata affettuosa.
Grazie a mamma e papà per avermi supportata e per aver creduto in me, sempre.
Grazie ad Alex, per essere rimasto fino alla fine.

“There is nothing like looking, if you want to find something. You certainly usually find
something, if you look, but it is not always quite the something you were after.”
J.R.R. Tolkien
LIST OF ABBREVIATIONS

ABC
ATP binding cassette; 15
ACh
Acetylcholine; 61
ADP
Adenosine diphosphate; 15
AOX
Alcohol oxidase promoter; 78
ATP
Adenosine triphosphate; 15
cAMP
Cyclic adenosine monophosphate; 59
CFTR
Cystic fibrosis transmembrane conductance regulator; 17
COPI
Coat protein complex I; 34
CTD
Cytoplasmic domain; 33
D2
L
Long Dopaminergic receptor 2; 51
DAG
Diacylglycerol; 57
DDM
N-dodecyl β-D-maltopyranoside; 81
DEND
Developmental delay, epilepsy, and neonatal diabetes; 44
DEPC
Diethyl pyrocarbonate; 72
DNA
Deoxyribonucleic acid; 71
ECF
Energy-coupling factor; 21
ECL
Extracellular loop; 52
eGFP
Enhanced green fluorescent protein; 79
EGTA
Ethylene glycol tetra-acetic acid; 87
ER
Endoplasmic reticulum; 16
GDP
Guanosine triphosphate; 40
GIRK
G-protein activated inward rectifying potassium channel; 32
GPCR
G protein-coupled receptor; 40
GRAFS
Glutamate, rhodopsin, adhesion, frizzled/taste2, secretin receptor classes; 53
HA
Human influenza hemagglutinin; 75
hOR
Human olfactory receptor; 51
ICCRs
Ion channel-coupled receptor; 50
ICL
Intracellular loop; 52
IP3
Inositol trisphosphate; 57
IPTG
Isopropyl β-D-1-thiogalactopyranoside; 75
K-ATP
ATP-sensitive potassium channel; 15
KCO
Potassium channel opener; 48
Kir
Inwardly rectifying potassium channel; 15
LB
Luria-Bertani; 71
LC-CoA
long chain Co-enzyme A esters; 43
M2
Muscarinic receptor 2; 51
MAC
Metal affinity chromatography; 81
MNG-3
Maltose-neopentyl glycol; 81
mRNA
Messenger ribonucleic acid; 73
MRP
Multidrug resistance protein; 17
MSP
Membrane scaffold protein; 80
NBD
Nucleotide binding domain; 17
PBS
Phosphate-buffer saline; 82
PCR
Polymerase chain reaction; 78
P-gp
P-glycoprotein; 24
PHHI
Persistent hyperinsulinemic hypoglycaemia of infancy; 35
PIP
2
Phosphatidylinositol 4,5-bisphosphate; 42