Topic "Molecular Engineering"

A few words

Expertise in the engineering of redox membrane proteins

The I2M team is also recognized for its know-how in the field of protein engineering, in particular cytochromes P450 and their electron donors. We have developed original approaches for the creation of combinatorial libraries and functional mapping of modified enzymes. The engineering methods we use target either individual enzymes (modification of natural enzyme activities, engineering of active or recognition sites) or macromolecular assemblies (formation of redox complexes, formation of the shell of bacterial microcompartments, assemblies of individual enzymes). Molecular engineering is perfectly integrated with the metabolic engineering theme because both pursue the same objective: to obtain efficient synthetic metabolic systems. The enzymatic models used are oxidases (cytochromes P450), terpene synthetases and enzymes of the carotenoid production pathway (desaturases, cyclases, dioxygenases, etc.).

Ingenierie Proteines I2M

Ongoing projects

Project

Bacterial Microcompartments

Synthetic continuously demands the development of novel biotechnology tools to improve the efficiency of metabolic cascades engineered within given microorganisms. Two major strategies for reaching this objective are the spatial colocalization of the designed enzymes, and the optimization of their relative orientation. In that sense, our first intention is to learn from nature, namely contributing to our understanding of bacterial micro-compartments (BMC) function. BMC are macromolecular assemblies that naturally encapsulate enzymes that participate to various metabolic processes. We investigate by diverse biophysical means the mechanism that leads to BMC shell assembly. The goal of the second axis of this project is to establish the precise composition of natural assembly units for then engineering new hexameric platforms.

Project

Combination of linker sequences on the activity of artificial fusion proteins.

 

Project

Interdomain dynamics in a multidomain oxidoreductase

Dans l’organisme, les médicaments subissent un métabolisme spécifique qui s’effectue principalement par les enzymes P450. Une seule flavoenzyme, la NADPH-cytochrome P450 réductase (CPR), donne les électrons nécessaires aux P450s pour leur catalyse. Jusqu’à présent, la plupart des variations interindividuelles de ce métabolisme ont été étudiées sur les polymorphismes du P450. Les modifications de l’équilibre conformationnel de la CPR, causées soit par des mutations, soit par l’interaction avec de petites molécules, peuvent augmenter son affinité pour un P450 spécifique, au détriment des autres. L’objectif de ce projet est de déchiffrer les facteurs (membrane du reticulum endoplasmique, état d’oxydation et variantes polymorphes de la CPR) qui contrôlent les modifications structurales et la dynamique de la CPR ancrée dans la membrane et leur implication dans le métabolisme des médicaments et les déficiences fonctionnelles potentielles. Ce projet vise à fournir de nouveaux outils pour décrypter les facteurs moléculaires modulant le paysage dynamique de la CPR et la spécificité des interactions dans les complexes CPR/P450.

Projet AlphaFold2

AlphaFold and protein complexes

AlphaFold2, an AI deep-learning system developed by DeepMind surpasses in accuracy all previous computing attempts for protein structure predictions. Recent versions were successfully extended to   modeling of protein complexes including in some instances the potential to address large scale conformational changes involved in catalysis. DeepMind and EMBL’s European Bioinformatics Institute (EMBL-EBI) have partnered to create AlphaFold Protein Structure DataBase (ADB) that covers the complete human and 47 other key organisms (mouse, yeast, Arabidopsis, etc.), as well as the manually curated UniProt database. As of April 2022, AlphaFold DB provides open access to 992,316 protein 3D structure entries, most of them with no experimental structure available. We are using AlphaFold2_advanced version to look at the structures of complexes between P450 enzymes and P450-associated redox proteins and to their alternative geometries.