Enzymes: catalyse, inhibition et méthodologie en cristallographie
Our team aims to study in fine detail the structural basis of ligand binding to macromolecules involved in important diseases (targets) and the corresponding structure activity relationships (SAR), in order to gather information for the “hit to lead” discovery process. We determine the crystallographic structures of proteins and macromolecular complexes, in the presence and in the absence of natural or synthetic ligands. We develop new methodologies, in particular for high resolution X-Ray and neutron diffraction, to improve the quality of the structures of ligand-target complexes. We collaborate with several national and international groups for the production of ligands and for the modeling of ligand binding, which we use to complement the experimental structural information. In summary we apply an integrated approach to Experimental Structure Based Drug Design.
In structural biology our expertise covers cloning, protein expression in prokaryotic and eukaryotic cells, large scale protein production, purification, protein crystallization and protein crystallography. The latter is the main tool to visualize macromolecular complexes.
We develop special crystallographic methods based both on high resolution X-Rays (PX) and on Neutrons (NPX) diffraction, leading to very accurate models including the observation of protonation states and hydration. This exceptional level of joint information is then used to explain very precisely the mechanisms of action and inhibition. We also study in detail the thermodynamics of binding. In this sense we apply several biochemical and biophysical techniques, including Isothermal Titration Calorimetry (ITC), Mass Spectrometry (MS), Surface Plasmon Resonance (SPR) and analytical ultra-centrifugation.
As a significant example, we have studied one protein, human Aldose Reductase, at the highest resolution ever obtained so far with X-Ray Crystallography (0.66 Å) for a medium size enzyme (36 kDa). We have used this information for the development of pharmaceutical leads. One of these leads (Fidarestat) is currently in phase III of clinical trials.
Projets en cours
We are currently studying several pharmaceutical targets, the most important ones being:
1) Aldo-Keto reductases. We have a large experience in human Aldose Reductase, an enzyme involved in Diabetes complications. We have extended this project to AKR1B10, an enzyme involved in cancer tumor growth.
2) Fatty acid binding proteins (FABPs). FABPs modulate intracellular lipid homeostasis by regulating FA transport in the nuclear and extra-nuclear compartments of the cell; in this process, they also impact systemic energy homeostasis. Cardiac FABPs are involved in heart function, and also an important marker of cardiac infarction.
Other targets under study are:
3) Human Arginase
4) Acidic mammalian chitinase
1. X- Ray High resolution data collection and refinement at room and cryo temperatures.
Andrzej Joachimiak, Synchrotron APS, USA
Vincent Olieric and Takashi Tomizaki, Synchrotron SLS, Switzerland
Christophe Mueller-Dickmann and Alexandre Popov, Synchrotron ESRF, France
Tatiana Petrova, IMPB institut, Russia
2. Quantum modeling
Raul Cachau, National Cancer Institute, Frederick, USA
Oscar Ventura, Universidad de la Republica, Uruguay
3. Neutron protein crystallography
Michael Haertlein and Matthew Blakeley, ILL, Grenoble, France
Nobuo Niimura, Ibaraki University, Japon
4. Fatty acid binding proteins
Eduardo Howard, IFLYSIB institut, La Plata, Argentina
Luis Beaugé, Universidad de Cordoba, Argentina
5. Studies of Aldo-Keto reductase complexes and inhibitors
Xavier Parés, Universitat Autònoma de Barcelona, Spain
Gerhard Klebe, Philipps-Universität Marburg, Germany
Angel de Lera, Universidad de Vigo, Spain
6. Protein Stability
Javier Santos, Universidad de Buenos Aires, Argentina
7. Mass Spectrometry
Alain Van Dorsselaer and Sarah Sanglier, IPHC, Strasbourg, France
8. Surface Plasmon Resonance
Danièle Altschuh, ESBS, Illkirch/Strasbourg, France
Sci Rep 9 février 2017 ; 7:42343 .
Chem Biol Interact 1 février 2017 .
Biochim Biophys Acta Jun 2016 ; 1864:655-66 .
ACS Chem Biol 21 octobre 2016 ; 11:2693-2705 .
PLoS One 25 avril 2016 ; 11:e0154190 .
High-resolution neutron and X-ray diffraction room-temperature studies of an H-FABP-oleic acid complex: study of the internal water cluster and ligand binding by a transferred multipolar electron-density distribution.
IUCrJ 16 janvier 2016 ; 3:115-126 .
Chemistry 4 avril 2016 ; 22:5151-5 .
Nature 28 janvier 2016 ; 529:541-5 .
Proteins Jan 2016 ; 84:129-42 .
J Biol Inorg Chem Jun 2015 ; 20:653-64 .
Sujets de thèse