Structural insights on the pamoic acid and the 8 kDa domain of DNA polymerase beta complex: Towards the design of higher-affinity inhibitors

<p>Abstract</p> <p>Background</p> <p>DNA polymerase beta (pol beta), the error-prone DNA polymerase of single-stranded DNA break repair as well as base excision repair pathways, is overexpressed in several tumors and takes part in chemotherapeutic agent resistance, like that of cisplatin, through translesion synthesis. For this reason pol beta has become a therapeutic target. Several inhibitors have been identified, but none of them presents a sufficient affinity and specificity to become a drug. The fragment-based inhibitor design allows an important improvement in affinity of small molecules. The initial and critical step for setting up the fragment-based strategy consists in the identification and structural characterization of the first fragment bound to the target.</p> <p>Results</p> <p>We have performed docking studies of pamoic acid, a 9 micromolar pol beta inhibitor, and found that it binds in a single pocket at the surface of the 8 kDa domain of pol beta. However, docking studies provided five possible conformations for pamoic acid in this site. NMR experiments were performed on the complex to select a single conformation among the five retained. Chemical Shift Mapping data confirmed pamoic acid binding site found by docking while NOESY and saturation transfer experiments provided distances between pairs of protons from the pamoic acid and those of the 8 kDa domain that allowed the identification of the correct conformation.</p> <p>Conclusion</p> <p>Combining NMR experiments on the complex with docking results allowed us to build a three-dimensional structural model. This model serves as the starting point for further structural studies aimed at improving the affinity of pamoic acid for binding to DNA polymerase beta.</p

Structural insights into chaperone addiction of toxin-antitoxin systems

International audienceSecB chaperones assist protein export by binding both unfolded proteins and the SecA motor. Certain SecB homologs can also control toxin-antitoxin (TA) systems known to modulate bacterial growth in response to stress. In such TA-chaperone (TAC) systems, SecB assists the folding and prevents degradation of the antitoxin, thus facilitating toxin inhibition. Chaperone dependency is conferred by a C-terminal extension in the antitoxin known as chaperone addiction (ChAD) sequence, which makes the antitoxin aggregation-prone and prevents toxin inhibition. Using TAC of Mycobacterium tuberculosis, we present the structure of a SecB-like chaperone bound to its ChAD peptide. We find differences in the binding interfaces when compared to SecB–SecA or SecB-preprotein complexes, and show that the antitoxin can reach a functional form while bound to the chaperone. This work reveals how chaperones can use discrete surface binding regions to accommodate different clients or partners and thereby expand their substrate repertoire and functions

Structural determinants of specific DNA-recognition by the THAP zinc finger

Human THAP1 is the prototype of a large family of cellular factors sharing an original THAP zinc-finger motif responsible for DNA binding. Human THAP1 regulates endothelial cell proliferation and G1/S cell-cycle progression, through modulation of pRb/E2F cell-cycle target genes including rrm1. Recently, mutations in THAP1 have been found to cause DYT6 primary torsion dystonia, a human neurological disease. We report here the first 3D structure of the complex formed by the DNA-binding domain of THAP1 and its specific DNA target (THABS) found within the rrm1 target gene. The THAP zinc finger uses its double-stranded β-sheet to fill the DNA major groove and provides a unique combination of contacts from the β-sheet, the N-terminal tail and surrounding loops toward the five invariant base pairs of the THABS sequence. Our studies reveal unprecedented insights into the specific DNA recognition mechanisms within this large family of proteins controlling cell proliferation, cell cycle and pluripotency

The CADM1 tumor suppressor gene is a major candidate gene in MDS with deletion of the long arm of chromosome 11.

Myelodysplastic syndromes (MDS) represent a heterogeneous group of clonal hematopoietic stem cell disorders characterized by ineffective hematopoiesis leading to peripheral cytopenias and in a substantial proportion of cases to acute myeloid leukemia. The deletion of the long arm of chromosome 11, del(11q), is a rare but recurrent clonal event in MDS. Here, we detail the largest series of 113 cases of MDS and myelodysplastic syndromes/myeloproliferative neoplasms (MDS/MPN) harboring a del(11q) analyzed at clinical, cytological, cytogenetic, and molecular levels. Female predominance, a survival prognosis similar to other MDS, a low monocyte count, and dysmegakaryopoiesis were the specific clinical and cytological features of del(11q) MDS. In most cases, del(11q) was isolated, primary and interstitial encompassing the 11q22-23 region containing ATM, KMT2A, and CBL genes. The common deleted region at 11q23.2 is centered on an intergenic region between CADM1 (also known as Tumor Suppressor in Lung Cancer 1) and NXPE2. CADM1 was expressed in all myeloid cells analyzed in contrast to NXPE2. At the functional level, the deletion of Cadm1 in murine Lineage-Sca1+Kit+ cells modifies the lymphoid-to-myeloid ratio in bone marrow, although not altering their multilineage hematopoietic reconstitution potential after syngenic transplantation. Together with the frequent simultaneous deletions of KMT2A, ATM, and CBL and mutations of ASXL1, SF3B1, and CBL, we show that CADM1 may be important in the physiopathology of the del(11q) MDS, extending its role as tumor-suppressor gene from solid tumors to hematopoietic malignancies

Elaboration de céramiques CaCu3Ti4O12 à haute constante diélectrique et réalisation de condensateurs "tout oxyde" par ablation laser pulsé

The oxide CaCu3Ti4O12 (CCTO) shows a very high permittivity (~ 105) in a large range of temperature (100-60K) and frequencies (10Hz-1MHz). A part of this study is an atttempt to deposit "all oxide" capacitors based on CCTO using the laser on CCTO using the laser ablation technique. At first, epitaxial films of CCTO were deposited on single crystal LaA1O3(100) (LAO) after optimisation of the target sintering. Capacitors LSNO/CCTO/LSNO/LAO with LaSrNiO4 (LSNO) as electrodes were then structured in-situ by shadow-masking. In an attempt to reduce the diffusion of La and Cu and to improve the quality interface with electrode, LSNO was replaced by CaCu3Ru4O12 which was isostructural of CCTO. In a second part, CCTO in the form of pure and doped powder and thin film was studied by electron spin resonance. New results are important to understand the mechanisms giving rise to the high permittivity value.L'oxyde CaCu3Ti4O12 (CCTO) présente une permittivité élevée (~ 105) dans une large gamme de température (100-600K) et de fréquence (10HZ-1MHz). La première partie de cette étude est une tentative de dépôt de condensateurs "tout oxyde" à base de CCTO par ablation laser. Des fims épitaxiés de CCTO sur monocristal LaA1O3 (001) ont été obtenus après optimisation du frittage des cibles. LaSrNiO4 (LSNO) a servi d'électrode pour des condensateurs LSNO/CCTO/LSNO/LAO, structurés in situ par "shadow-masking". Afin de réduire la diffusion du lanthane et de cuivre et d'améliorer la qualité des interfaces avec l'électrode, une nouvelle électrode CaCu3Ru4O12 isostructural de CCTO a été développée. En parallèle, CCTO sous forme de poudre pure et dopée et de film a été étudié par RPE. Les résultats originaux sont d'importance pour la compréhension des mécanismes à l'origine des propriétés exceptionnelles de CCTO.TOURS-BU Sciences Pharmacie (372612104) / SudocPARIS-BIUP (751062107) / SudocSudocFranceF

Déterminants structuraux de la reconnaissance spécifique de l'ADN par le domaine THAP de hTHAP1 et implications dans la dystonie DYT6

TOULOUSE3-BU Sciences (315552104) / SudocPARIS-BIUP (751062107) / SudocSudocFranceF

Le domaine THAP de THAP1 (structure par RMN en solution et interaction avec l'ADN)

TOULOUSE3-BU Sciences (315552104) / SudocSudocFranceF

Complexation Preferences of Dynamic Constitutional Frameworks as Adaptive Gene Vectors

The growing applications of therapeutic nucleic acids requires the concomitant development of vectors that are optimized to complex one type of nucleic acid, forming nanoparticles suitable for further trafficking and delivery. While fine-tuning a vector by molecular engineering to obtain a particular nanoscale organization at the nanoparticle level can be a challenging endeavor, we turned the situation around and instead screened the complexation preferences of dynamic constitutional frameworks toward different types of DNAs. Dynamic constitutional frameworks (DCF) are recently-identified vectors by our group that can be prepared in a versatile manner through dynamic covalent chemistry. Herein, we designed and synthesized 40 new DCFs that vary in hydrophilic/hydrophobic balance, number of cationic headgroups. The results of DNA complexation obtained through gel electrophoresis and fluorescent displacement assays reveal binding preferences of different DCFs toward different DNAs. The formation of compact spherical architectures with an optimal diameter of 100–200 nm suggests that condensation into nanoparticles is more effective for longer PEG chains and PEI groups that induce a better binding performance in the presence of DNA targets

Switching Multivalent DNA Complexation using Metal-Controlled Cationic Supramolecular Self-Assemblies

International audienc