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Breakthrough a glycyl radical enzyme long range activation mechanism

Project description

Revealing the activation of glycyl radical enzymes

Glycyl radical enzymes (GREs) catalyse a wealth of biochemical reactions in anaerobic microorganisms and in the human body. They are responsible for many types of radical-based chemical reactions in diverse metabolic pathways: DNA synthesis, acid fermentation, and the anaerobic metabolism of pollutants, for instance. Characterising their mechanism of activation which requires the formation of a protein-protein activation complex with small GRE-activating enzymes (AEs) is quite challenging due to their sensitivity to the presence of oxygen. With the support of the Marie Skłodowska-Curie Actions programme, the GREAM project aims to combine biochemical and biophysical studies assisted by cryo-electron microscopy (EM) and X-ray crystallography to solve the anaerobic cryo-EM structure of an important GRE:AE holocomplex.

Objective

Glycyl radical enzymes (GREs) are one of the most prominent biological catalysts in both strict and facultative anaerobes. They are responsible for a broad range of radical-based chemistry and catalyse reactions involved in diverse metabolic pathways such as acid fermentation, DNA synthesis and the anaerobic metabolism of pollutants. GREs are activated by members of the S-adenosylmethionine (SAM) radical family. Activation of GREs requires the formation of a protein-protein activation complex with the small GRE-AE. All known structures of GREs show that a major conformational change very likely occurs during the glycyl radical generation upon interaction with the GRE-AE. Only a structure of a GRE:AE holocomplex would shed light on the unprecedented conformational changes involved in GRE activation. Glycyl radicals and iron-sulfur clusters are extremely oxygen sensitive, rendering the proteins challenging to work with.

GREAM will narrow a significant knowledge gap by studying the anaerobic class III RNR natural fusion (NrdD:NrdG). In order to study the mechanism of activation, GREAM will use an integrative research approach combining biochemical, biophysical and cryo-EM assisted by X-ray crystallography.

With my experience in anaerobic protein characterisation and crystallisation, I will join the Fontecave lab to build up on their experience in spectroscopic characterisation of oxygen-sensitive protein complexes and achieve the first aim or GREAM. During a secondment in the Logan lab, who are expert in structural characterisation of aRNRs and have a long lasting collaboration with the host, I will solve the anaerobic cryo-EM structure of the NrdD:NrdG holocomplex and achieve the final goal of GREAM.

This newly established international collaboration will permit combining integrative approaches to successfully answer all GREAMs questions and allow me to acquire invaluable experience in my future career.

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HORIZON-TMA-MSCA-PF-EF - HORIZON TMA MSCA Postdoctoral Fellowships - European Fellowships

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Call for proposal

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(opens in new window) HORIZON-MSCA-2023-PF-01

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Coordinator

COLLEGE DE FRANCE
Net EU contribution

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€ 211 754,88
Address
PLACE MARCELIN BERTHELOT 11
75005 PARIS
France

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Region
Ile-de-France Ile-de-France Paris
Activity type
Higher or Secondary Education Establishments
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Total cost

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