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Protein dynamics with fast magic-angle spinning NMR

Date
mer 09 déc 2020
Horaires

15h00

Lieu(x)
CRMN

 5 rue de la Doua - Villeurbanne

Intervenant(s)

Soutenance de Mme BONACCORSI Marta sous la Direction de M. PINTACUDA Guido 

Langue(s) des interventions

Description générale

The aim of my thesis is to develop Magic-Angle Spinning (MAS) NMR to characterize structure and dynamics in complex biological samples, with a particular focus on the role of metal ions in enzymes and channels.
MAS NMR is a powerful technique that allows to extract atomic level information, characterize broad timescales of motions, and investigate functional states in native-like sample conditions, a particularly important requirement e.g. for transmembrane proteins in lipid bilayers. Nonetheless, a number of bottlenecks prevents its widespread application in structural biology.
In my work I developed and applied tailored techniques based on high magnetic fields (800 MHz and 1 GHz) and MAS probes with sub-mm diameter rotors spinning at rates above 100 kHz, which contributed to push forward the capability of this technique: i) by enlarging the molecular size of the systems that can be investigated with site specificity; ii) by reducing the requirements in terms of isotopic labeling, notably deuteration; iii) by speeding up the tedious processes of resonance assignment and acquisition of dynamical parameters; iv) by enriching the palette of measurable parameters connected to dynamics.
All along this thesis, the methods were first benchmarked on microcrystalline samples of the model domain GB1, then applied to Cu,Zn-superoxide dismutase (a dimeric 2x16 kDa Cu metalloenzyme) in functional microcrystalline form, as well as to two transmembrane channels reconstituted in lipid bilayers, bacterial CorA (a pentameric 5x40 kDa cation channel) and human Aqp-1 (tetrameric 4x25 kDa aquaporin-1 water channel). The data obtained shed new light on the relation between internal dynamics and function. 
 

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