Group of Marc Delarue

Projects (until 2009)

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puce  1. X-Ray crystallography

All these structures have benefitted from excellent X-ray data collection facilities in ESRF, Grenoble.

puce  1.1 X-Ray crystallography and Drug Design

puce TMP kinase from M. tuberculosis.

TMPK

The structure of thymidylate kinase TMPK which is essential for nucleotide metabolism has been solved at 1.95 Å resolution in the presence of TMP. We use this structural information to design potent new inhibitors of this target. The enzyme is active in the crystal state (see D. Bourgeois web site).

The TMPK project is a collaboration with H. Munier-Lehmann and A.M. Gilles in O. Barzu's lab (Laboratoire de Chimie Structurale des Macromolecules).

TMPK


puce 6PGL from T. brucei
6PGL

The Trypanosoma brucei 6-phosphogluconolactonase has been crystallized and solved at 2.1 Angstrom resolution. It is a member of the Pentose-Phosphate-Pathway (PPP) and is a good candidate for drug design against T. brucei.

This work is a collaboration with Veronique Stoven (Unite de BioInformatique Structurale) and was made possible thanks to both PT5 (P. Beguin) and PT6 (A. Haouz).

6PGL


puce  1.2 Structural Molecular Biology and DNA polymerases

puce Murine Terminal desoxynucleotidyl transferase.
TdT

Tdt is a member of the pol X family; it is responsible for the random addition of nucleotides at the N regions of immunoglobulins and T-cell receptors genes at the V(D)J junctions, during somatic recombination, thereby contributing to the generation of the diversity of the immune response (Tdt). Crystals of the catalytic domain of TdT have been obtained in the lab and the structure has been solved by MIR methods at 2.35 Å resolution. Binary complexes with either the incoming dNTP or the oligonucleotide primer have also been solved and analysed. A number of important biological implications of the structure have been found.

The TdT project is a collaboration with J.B. Boulé and C. Papanicolaou in F. Rougeon's lab (Unité de Génétique et de Biochimie du Développement).

TdT


puce  Human Pol &mu: see here for our latest results published in Nucleic Acids Res. (2009).
TdT

We ask the question of the molecular origin of the difference in activity (phenotype) of pol mu and Tdt, knowing that they share 42% of sequence identity. Using systematic mutations in the Loop1, SDR1 and SDR2 regions we managed each time to transform a template-independent Tdt into a template-dependent DNA polymerase by just a single-point mutation.

TdT

puce  DNA Pol X from bacteria.
TdT

We reinvestigate the structural dynamics of the respective orientation of domains in the polymerase (palm, fingers and thumb) and PHP functions in bacterial polX, using both SAXS and crystallography.

TdT

puce  DNA PolB from Archaea: this is the work of Jerome Gouge (PhD student), in collaboration with Ghislaine Henneke (IFREMER, Brest).
TdT

We use this system to investigate how the replication fork is stalled in archaea when an unusual base (dU, dH) is encountered 4 or 5 bases upstream the primer/template junction. Our results are in favor of a kinetic proofreading mechanism.

TdT

puce  1.3 Bacterial homologues of pentameric Ligand-gated Ion channels, in collaboration with P.J. Corringer (Dept Neuroscience, IP).

TdT

We solved the structure at 2.9 Angstrom resolution of a member of the Cys-Loop receptors family, in an open conformation. See Ref1, Ref2, Ref3. We are specially interested in the permeation process as well as the modulation of agonists activity by alcohol and/or anesthetics.

TdT


Publications


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puce  2. Mean-Field Optimization techniques in molecular modelling and crystallography


puce  2.1 Side chain prediction, homology modelling and sequence design.

Mean Field optimization can be used to "decorate" very rapidly a given protein backbone with any desired protein sequence. Compared to other similar techniques, MF optimization is very fast ; it was originally developed together with P. Koehl. The output of the program are the coordinates of all the atoms of the model.

Four main applications are available :


Publications


puce  2.2 Phase Refinement in Crystallography (Methodology)

Mean Field theory has been used to recast in a single formalism the problem of phase optimization and phase combination, generalizing the approach of Blow and Crick (1959) and Sim (1960) to treat rigourously density modification techniques in the presence of an experimentally derived phase probability distribution function in the same unifying theoretical framework.
In effect, a new statistical thermodynamics theory of crystallographic refinement has been set up. Because structure factors are complex numbers, field theoretic methods had to be used to calculate the partition function Z of the system, from which the free energy can then easily be derived. It bears strong resemble to maximum likelihood formalism but there are subtle differences.
The "best" maps are those that minimize the free energy. Their use should increase the efficiency of automatic model building programs (Arp/wArp) in structural genomics projects. Click here to find out more about the derived formula giving the generalized figure-of-merit and mean-phase FOM and mean-phase for different cases of density modification (use of a partial model, solvent flattening and Sayre formula) which are imposed as energy functions in reciprocal space: definitions of Yk for these three different cases.


The formalism bears strong resemblance with maximum likelihood, but can be readily interpreted in terms of usual thermodynamic functions such as free energy, entropy etc...

Publications

puce  2.3 Variations on Poisson-Boltzmann equation

We use Mean Field formalism to write down and solve variations of the Poisson-Boltzmann equation applied to proteins in different situations (finite size of the free ions...). We are especially interested in the case where the solvent is represented as an assembly of dipoles of variable density.
Eventually, this should have applications in drug design methodology, because it gives the solvation energy that is needed to estimate the free energy of binding of a ligand onto a macromolecule (protein).

Go to PDB_Hydro web site for a recent implementation of our Poisson-Boltzmann-Langevin model which treats the solvent as an assembly of self-optimised orientable dipoles of variable density.

N.B. This site also contains a number of programs to handle, repair and refine protein coordinates (PDB) files. This is our web site for our own computational structural biology on-line tools.

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puce  3. Normal Modes and the Elastic Network Model: a simplified approach to structural transitions in macromolecules


TdT

We discovered that Normal Modes are extremely good to describe structural transitions in polymerases.
Since then, we systematically use NMA for various applications of Structural Mechanics to proteins.
3.1 Click here NMA to go to main menu for Normal Modes Analysis, which allows for directly calculating and visualizing low-frequency Normal Modes of virtually any macromolecule on line, given its coordinates.
This also includes the possibility to refine macromolecular models against experimental X-Ray diffraction data and/or Electron Microscopy envelopes.
3.2 It can also generate plausible realistic trajectories between two conformations of the same macromolecule (much better than morphing!). Recently, an entirely new method based on the minimisation of Action along the trajectory was described and made available on the following web site MinActionPath. This is a collaboration with J. Franklin (Reed College), S. Doniach (Stanford) and P. Koehl (UC Davis).



Publications




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