Factor VIII-von Willebrand Factor Complex Inhibits Osteoclastogenesis and Controls Cell Survival

Factor VIII-von Willebrand factor (FVIII·vWF) complex, a molecule involved in coagulation, can be physically associated with osteoprotegerin (OPG). OPG is an anti-osteoclastic protein and a soluble receptor for the proapoptotic protein TRAIL (tumor necrosis factor-related apoptosis-inducing ligand), suggesting a potential role of FVIII·vWF complex in bone and cancer biology. We, thus, assessed the effects of FVIII·vWF complex on osteoclastogenesis and cell survival. We first evidenced that FVIII·vWF complex inhibited RANKL-induced osteoclastogenesis and enhanced the inhibitory effect of OPG. Interestingly, we revealed by surface plasmon resonance that FVIII·vWF complex bound to RANKL, whereas recombinant FVIII and vWF did not. By modeling, we showed that the OPG binding domain to the A1 domain of vWF was closely located and partially overlapped to its binding site to RANKL. Then, we demonstrated that FVIII·vWF complex cancelled the inhibitory activity of OPG on TRAIL-induced apoptosis and characterized interactions between these molecules. The present work evidenced a direct activity of FVIII·vWF complex on osteoclasts and on induced cell apoptosis, pointing out its potential involvement in physiological bone remodeling or in bone damages associated with severe hemophilia and cancer development

Etude par modélisation de dynamique moléculaire et spectroscopie RMN des déformations induites par la coordination du cisplatine sur l'ADN

Cisplatin (cis-diammin dichloro platin) is one of the most widely used anti cancer chemotherapy drug. Its anticancer activity has been rst described in 1965 by B. Rosenberg. Since then, many studies have been undertaken to decipher its mechanism of action. From an original approach linking molecular dynamics simulation and experimental studies, my PhD allowed a dynamic description of a platinated adduct on the sequence 5'-GCCG*G*GTCGC-3' / 5'-GCGACCCGGC-3' (where G* is a platinated guanine). The resulting structure has been studied previously in our laboratory on the identical sequence but with an A-T base pair on the 3' side of the platinated guanines (subsequently named G*G*A). Therefore we studied the structural implications of an A to G substition in the 3' region on the adduct structure. This is the fi rst structural study on a cisplatin adduct containing three adjacent guanines. GGG sequences and Gn (n>= 3) are known to be favored targets for platin (II) but their adducts have not been studied by NMR since their puri cation are problematic. This G*G*G NMR study has been compared to its dynamics simulation, where the parm 98 force field has been speci cally optimized for taking into account the platin coordination sphere. The parametrisation has been cross-validated by the NMR experiment. To calculate the proportion of BI/BII DNA substates, a novel method has been set up. Four distances, namely H2'(n)-H8(n), H1'(n)-H6/8(n+1), H2'(n)-H6/8(n+1) and H2''(n)-H6/8(n+1) allow to discriminate between the two substates and to calculate the proportion of each substate. The force fi eld optimizations and the BI/BII DNA substates discrimination method allowed a precise description of the cisplatin-DNA adduct, allowing to study one possible way of anti-cancer e ciency. The complex cisplatin-DNA-Protein (notably LEF I- Lymphoïd Enhanced Factor I) is a plausible candidate to explain why the cancerous cell elects apoptosis or tumor repair. The protein-DNA (without cisplatin) ensemble simulation has allowed to describe the recognition path of the DNA deformation by the protein and the implication of a water molecule in the recognition mechanism. All these studies on cisplatin-DNA allowed us to get enough knowledge on the resulting deformations. Therefore, we then studied the structural deformation induced by another platin-related complex, the pyrazolate-bis-platine. This complex has been designed to obtain a small DNA deformation, in order to get a different response than the one of cisplatin. The molecular dynamics simulation has indicated it induces a smaller kink and a different global deformation than cisplatin thus proving a di erent recognition mode. Since it is known by in vitro studies that the pyrazolate-bis-platine owns an anti-tumoral activity, the dynamics simulations suggest its cellular mechanism is different than the one of cisplatin. These PhD studies have allowed a better understanding of DNA-cisplatin, DNAcisplatin-protein dynamic description and to improve the de nition of platinated complex for force fi elds parm 98 (and parm 99) of the molecular dynamics simulation suite AMBER.Le cisplatine (ou cis-diammine, dichloro-platine) est l'un des composés chimiques les plus utilisés actuellement en chimiothérapie anticancéreuse. Depuis la description de ses propriétés anticancéreuses par B. Rosenberg en 1965 de nombreux travaux ont été effectués a n de décrire le mécanisme d'action lui conférant ses propriétés antitumorales. A travers une approche originale couplant modélisation et travail expérimental, les recherches réalisées durant ma thèse ont permis d'élucider le comportement dynamique d'un adduit platiné sur la séquence 5'-GCCG*G*GTCGC-3' / 5'-GCGACCCGGC-3' (G* représente une guanine platinée). Cette structure a été comparée à celle de l'adduit ADN-cisplatine déterminée précédemment au laboratoire sur la séquence G*G*A. Nous avons ainsi étudié l'influence d'une guanine adjacente en 3' au pontage GG-Pt sur la structure de l'adduit. Il s'agit de la première étude structurale sur un adduit du cisplatine avec la séquence GGG. Même si l'a nité de la séquence GGG et des sites contenant Gn (n>= 3) pour le platine (II) est connue depuis longtemps, ses adduits avec le cisplatine n'ont pas encore été étudiés par RMN à cause des problèmes posés par leur puri cation. Cette étude de l'adduit G*G*G par RMN a été confrontée à la description dynamique de cet adduit, calculée par simulation. La paramétrisation du champ de force parm 98 a été spéci quement a née pour mieux décrire l'environnement de l'atome du platine. La confrontation entre la simulation de l'adduit G*G*G-Pt et les données issues de l'étude par RMN a permis de valider notre paramétrisation. Pour déterminer les proportions précises des sous-états BI et BII de l'ADN, une méthode novatrice a été mise au point. Celle-ci est basée sur la combinaison de quatre distances inter-protons H2'(n)-H8(n), H1'(n)-H6/8(n+1), H2'(n)-H6/8(n+1) et H2''(n)-H6/8(n+1) qui permet de discriminer les deux sous-conformations. Ces améliorations du champ de force et de la méthode de détermination des sous-conformations BI/BII ont permis la description ne du comportement de l'adduit couplé à l'ADN, ce qui nous a servi pour étudier le mécanisme anti-tumoral du cisplatine. En effet la reconnaissance de l'ensemble cisplatine-ADN par une protéine (Lymhoïd Enhanced Factor I - LEF I) pourrait activer les voies métaboliques de la cellule cancéreuse conduisant vers l'apoptose ou vers la réparation de la tumeur. La simulation de l'ensemble ADN-protéine (sans cisplatine) a permis de présenter le mode de reconnaissance de la protéine sur la déformation ainsi que la mise en évidence de l'implication d'une molécule d'eau dans celui-ci. Les études sur le cisplatine fixé sur son ADN cible nous ayant apporté de nombreuses connaissances sur les déformations engendrées, nous avons validé la déformation structurale formée par un autre complexe de platine, le pyrazolato-bis-platine. Ce composé a été conçu de novo pour induire une déformation faible de son ADN cible a n de provoquer une réponse cellulaire différente de celle engendrée par le cisplatine. La simulation a indiqué que ce complexe induit une faible courbure de l'ADN et une déformation globale différente de celle du cisplatine, ce qui exclut le même mode de reconnaissance. Comme expérimentalement le complexe pyrazolato possède une activité anti-tumorale, les simulations effectuées suggèrent donc que le mode d'action au niveau cellulaire est différent du cisplatine. Les travaux réalisés lors de ma thèse ont permis d'améliorer la compréhension des déformations ADN-cisplatine, ADN-cisplatine-protéine et de perfectionner la description des composés platinés dans le champ de force parm 98 (puis parm 99) du logiciel de modélisation moléculaire AMBER

Olfactory receptor structure prediction and classification of ligand-receptor affinities

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A Review of the Literature Organized Into a New Database: RHeference

International audienceHundreds of articles containing heterogeneous data describe D variants or add to the knowledge of known alleles. Data can be difficult to find despite existing online blood group resources and genetic and literature databases. We have developed a modern, elaborate database for D variants, thanks to an extensive literature search with meticulous curation of 387 peer-reviewed articles and 80 abstracts from major conferences and other sources. RHeference contains entries for 710 RHD alleles, 11 RHCE alleles, 30 phenotype descriptions (preventing data loss from historical sources), 35 partly characterized alleles, 3 haplotypes, and 16 miscellaneous entries. The entries include molecular, phenotypic, serological, alloimmunization, haplotype, geographical, and other data, detailed for each source. The main characteristics are summarized for each entry. The sources for all information are included and easily accessible through doi and PMID links. Overall, the database contains more than 10,000 individual pieces of data. We have set up the database architecture based on our previous expertise on database setup and biocuration for other topics, using modern technologies such as the Django framework, BioPython, Bootstrap, and Jquery. This architecture allows an easy access to data and enables simple and complex queries: combining multiple mutations, keywords, or any of the characteristics included in the database. RHeference provides a complement to existing resources and will continue to grow as our knowledge expands and new articles are published. The database url is http://www.rheference.org/

Prédiction de la structure des récepteurs olfactifs et classement des affinités

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Investigation of Phospholipase Cγ1 Interaction with SLP76 Using Molecular Modeling Methods for Identifying Novel Inhibitors

International audienceThe enzyme phospholipase C gamma 1 (PLCγ1) has been identified as a potential drug target of interest for various pathological conditions such as immune disorders, systemic lupus erythematosus, and cancers. Targeting its SH3 domain has been recognized as an efficient pharmacological approach for drug discovery against PLCγ1. Therefore, for the first time, a combination of various biophysical methods has been employed to shed light on the atomistic interactions between PLCγ1 and its known binding partners. Indeed, molecular modeling of PLCγ1 with SLP76 peptide and with previously reported inhibitors (ritonavir, anethole, daunorubicin, diflunisal, and rosiglitazone) facilitated the identification of the common critical residues (Gln805, Arg806, Asp808, Glu809, Asp825, Gly827, and Trp828) as well as the quantification of their interaction through binding energies calculations. These features are in agreement with previous experimental data. Such an in depth biophysical analysis of each complex provides an opportunity to identify new inhibitors through pharmacophore mapping, molecular docking and MD simulations. From such a systematic procedure, a total of seven compounds emerged as promising inhibitors, all characterized by a strong binding with PLCγ1 and a comparable or higher binding affinity to ritonavir (∆Gbind < −25 kcal/mol), one of the most potent inhibitor reported till no

Olfactory receptor structure prediction and classification of odor - receptor affinities

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Automatic modeling and prediction of G-coupled protein receptor-ligand complexes

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