Crystal structure of the phosphatidylethanolamine-binding protein from bovine brain: a novel structural class of phospholipid-binding proteins.

International audiencePhosphatidylethanolamine-binding protein (PEBP) is a basic protein found in numerous tissues from a wide range of species. The screening of gene and protein data banks defines a family of PEBP-related proteins that are present in a variety of organisms, including Drosophila and inferior eukaryotes. PEBP binds to phosphatidylethanolamine and nucleotides in vitro, but its biological function in vivo is not yet known. The expression of PEBP and related proteins seems to be correlated with development and cell morphogenesis, however. To obtain new insights into the PEBP family and its potential functions, we initiated a crystallographic study of bovine brain PEPB

Comparaison de deux affinements de la structure de la cyanite

La cyanite Al₂SiO₅ est triclinique et possède une pseudosymétrie orthorhombique. Ses traits essentiels ont été déterminés par Taylor (1929). Pour estimer la pseudosymétrie du champ cristallin au voisinage de l'aluminium il est nécessaire de connaître les coordonnées atomiques avec grande précision (de l'ordre de 0,01 Å). L'affinement de cette structure par la méthode des moindres carrés a été entrepris à partir des intensités de diffraction mesurées dans les conditions suivantes : — appareil enregistreur : goniomètre de Weissenberg — rayonnement utilisé : MoKα — partie du réseau réciproque enregistrée : réflexions hk0, hk7, hk8, h0l. Les résultats sont comparés à ceux obtenus par Burnham (1963). Cet auteur avait mesuré les intensités dans les conditions suivantes : — appareil enregistreur : diffractomètre — rayonnement utilisé : CuKα — partie du réseau réciproque enregistré : réflexions hkl. A la fin des deux affinements, les coordonnées atomiques sont très proches et les écarts types sont du même ordre de grandeur (σ = 0,004 Å).Rango Colette de, Tsoucaris Georges, Zelwer C., Devaux J. Comparaison de deux affinements de la structure de la cyanite. In: Bulletin de la Société française de Minéralogie et de Cristallographie, volume 89, 4, 1966. pp. 419-424

Comparaison de deux affinements de la structure de la cyanite

La cyanite Al₂SiO₅ est triclinique et possède une pseudosymétrie orthorhombique. Ses traits essentiels ont été déterminés par Taylor (1929). Pour estimer la pseudosymétrie du champ cristallin au voisinage de l'aluminium il est nécessaire de connaître les coordonnées atomiques avec grande précision (de l'ordre de 0,01 Å). L'affinement de cette structure par la méthode des moindres carrés a été entrepris à partir des intensités de diffraction mesurées dans les conditions suivantes : — appareil enregistreur : goniomètre de Weissenberg — rayonnement utilisé : MoKα — partie du réseau réciproque enregistrée : réflexions hk0, hk7, hk8, h0l. Les résultats sont comparés à ceux obtenus par Burnham (1963). Cet auteur avait mesuré les intensités dans les conditions suivantes : — appareil enregistreur : diffractomètre — rayonnement utilisé : CuKα — partie du réseau réciproque enregistré : réflexions hkl. A la fin des deux affinements, les coordonnées atomiques sont très proches et les écarts types sont du même ordre de grandeur (σ = 0,004 Å).Rango Colette de, Tsoucaris Georges, Zelwer C., Devaux J. Comparaison de deux affinements de la structure de la cyanite. In: Bulletin de la Société française de Minéralogie et de Cristallographie, volume 89, 4, 1966. pp. 419-424

Crystallization and preliminary X-ray diffraction analysis of the homodimeric form alpha2 of the HU protein from Escherichia coli.

International audienceThe homodimeric form alpha(2) of the Escherichia coli DNA-binding protein HU was crystallized by the hanging-drop vapour-diffusion method using PEG 4000 as a precipitant. The crystals belong to space group I222, with unit-cell parameters a = 31.09, b = 55.34, c = 117. 63 A, and contain one monomer per asymmetric unit. A full diffraction data set was collected to 2.3 A resolution on a conventional X-ray source. The molecular-replacement method, using the HU crystallographic model from Bacillus stearothermophilus as a starting point, gave a reliable solution for the rotation and translation functions

Structural basis for the recognition of the FapydG lesion (2,6-diamino-4-hydroxy-5-formamidopyrimidine) by Formamidopyrimidine-DNA glycosylase

Formamidopyrimidine-DNA glycosylase (Fpg) is a DNA repair enzyme that excises oxidized purines such as 7,8-dihydro-8-oxoguanine (8-oxoG) and 2,6-diamino-4-hydroxy-5-formamidopyrimidine (FapyG) from damaged DNA. Here, we report the crystal structure of the Fpg protein from Lactococcus lactis (LlFpg) bound to a carbocyclic FapydG (cFapydG)-containing DNA. The structure reveals that Fpg stabilizes the cFapydG nucleoside into an extrahelical conformation inside its substrate binding pocket. In contrast to the recognition of the 8-oxodG lesion, which is bound with the glycosidic bond in a syn conformation, the cFapydG lesion displays in the complex an anti conformation. Furthermore, Fpg establishes interactions with all the functional groups of the FapyG base lesion, which can be classified in two categories: (i) those specifying a purine-derived lesion (here a guanine) involved in the Watson-Crick face recognition of the lesion and probably contributing to an optimal orientation of the pyrimidine ring moiety in the binding pocket and (ii) those specifying the imidazole ring-opened moiety of FapyG and probably participating also in the rotameric selection of the FapydG nucleobase. These interactions involve strictly conserved Fpg residues and structural water molecules mediated interactions. The significant differences between the Fpg recognition modes of 8-oxodG and FapydG provide new insights into the Fpg substrate specificity

AP site structural determinants for Fpg specific recognition.

The binding of Escherichia coli and Lactococcus lactis Fapy-DNA glyosylase (Fpg) proteins to DNA containing either cyclic or non-cyclic abasic (AP) site analogs was investigated by electrophoretic mobility shift assay (EMSA) and by footprinting experiments. We showed that the reduced AP site is the best substrate analog for the E.coli and L.lactis enzymes ( K Dapp = 0.26 and 0.5 nM, respectively) as compared with the other analogs tested in this study ( K Dapp >2.8 nM). The 1,3-propanediol (Pr) residue-containing DNA seems to be the minimal AP site structure allowing a Fpg specific DNA binding, since the ethyleneglycol residue is not specifically bound by these enzymes. The newly described cyclopentanol residue is better recognized than tetrahydrofuran (for the E.coli Fpg, K Dapp = 2.9 and 25 nM, respectively). These results suggest that the hemiacetal form of the AP site is negatively discriminated by the Fpg protein suggesting a hydrogen bond between the C4'-hydroxyl group of the sugar and a Fpg residue. High-resolution hydroxyl radical footprinting using a duplex containing Pr shows that Fpg binds to six nucleotides on the strand containing the AP site and only the base opposite the lesion on the undamaged complementary strand. This comparative study provides new information about the molecular mechanism involved in the Fpg AP lyase activity

Crystal structure of a double-stranded DNA containing a cisplatin interstrand cross-link at 1.63 A resolution: hydration at the platinated site.

cis-diamminedichloroplatinum (II) (cisplatin) is a powerful anti-tumor drug whose target is cellular DNA. In the reaction between DNA and cisplatin, covalent intrastrand and interstrand cross-links (ICL) are formed. Two solution structures of the ICL have been published recently. In both models the double-helix is bent and unwound but with significantly different angle values. We solved the crystal structure at 100K of a double-stranded DNA decamer containing a single cisplatin ICL, using the anomalous scattering (MAD) of platinum as a unique source of phase information. We found 47 degrees for double-helix bending and 70 degrees for unwinding in agreement with previous electrophoretic assays. The crystals are stabilized by intermolecular contacts involving two cytosines extruded from the double-helix, one of which makes a triplet with a terminal G.C pair. The platinum coordination is nearly square and the platinum residue is embedded into a cage of nine water molecules linked to the cross-linked guanines, to the two amine groups, and to the phosphodiester backbone through other water molecules. This water molecule organization is discussed in relation with the chemical stability of the ICL