Preliminary X-ray analysis of a new crystal form of the Escherichia coli KDO8P synthase

3-Deoxy-d-manno-octulosonate 8-phosphate (KDO8P) synthase catalyzes the biosynthesis of an essential component of the lipopolysaccharide of all Gram-negative bacteria. The structure and mechanism of KDO8P synthase are being actively studied as this enzyme represents an important target for antibiotic therapy. The structure of the Escherichia coli KDO8P synthase in cubic crystals (space group I23) has recently been determined and the enzyme shown to be a tetramer of identical subunits. However, this information is challenged by biochemical studies, which suggest that the enzyme behaves in solution as a homotrimer. Here, the preparation and preliminary X-ray analysis of monoclinic crystals of KDO8P synthase are reported. The crystals belong to space group P21, with unit-cell parameters a≃ 50, b≃ 140, c≃ 74 Å, β≃ 105°. The structure of KDO8P synthase in the monoclinic crystal form was determined by molecular replacement, using as a search model one of the subunits of the enzyme in the cubic crystals. A tetramer of KDO8P synthase with 222 local symmetry is also present in the asymmetric unit of the P21 crystals, with a solvent content of 43%. The observation that the same quaternary structure of KDO8P synthase is observed in two different crystal forms belonging to distinct crystal systems (monoclinic and cubic) suggests that a tetramer is the native form of the enzyme

A rational approach to heavy-atom derivative screening

In order to overcome the difficulties associated with the ‘classical’ heavy-atom derivatization procedure, an attempt has been made to develop a rational crystal-free heavy-atom-derivative screening method and a quick-soak derivatization procedure which allows heavy-atom compound identification

La nuova Russia. Dibattito culturale e modello di società in costruzione

Una radiografia del dibattito culturale della Russia che illustra i tanti problemi con cui si deve confrontare la costruzione di una nuova società. Modernizzazione, tradizionalismo, nuovo corporativismo, privatizzazioni, nazionalismo e religione sono solo alcuni degli elementi che compongono un quadro ricco di conflittualità e foriero di grandi trasformazioni.- Indice #5- Radiografia del dibattito culturale: modernizzazione e tradizionalismo nello scontro tra gli “addetti ai lavori”, Ilja Levin #9- Partiti e società: evoluzione e prospettive della differenziazione politica ed ideologica nella Russia postsovietica, Kirill Kholodkovski #29- Dal “collettivismo” all’individualismo: l’uomo nella società deistituzionalizzata, Guerman Diliguenski #45- Gruppi direttivi regionali: trasformazione dei meccanismi organizzativi e relazionali del potere, Mikhail Afanasjev #63- L’”oligarchia” e la crisi in atto nel postcomunismo russo, Alexeij Zudin #81- Il nuovo corporativismo russo all’interno del contesto globale: il ruolo in via di cambiamento dell’élite settoriale, Sergei Peregudov #107- La formazione di nuovi mercati e lo Stato nella Russia postcomunista, Vadim Radaev #133- Stabilizzazione macroeconomica e mutamenti strutturali nell’economia nazionale russa: deindustrializzazione o sfondamento verso una società postindustriale?, Vladimir Mau e Irina Starodubrovskaja #163- Dopo la privatizzazione: alla ricerca di un modello ottimale di società, Viktor Studentsov #187- Fondamenta e principi dello Stato accentratore in Russia, Igor Pantin #217- Nazionalismo liberale o idea imperiale? La consapevolezza degli interessi e degli orientamenti nazionali in Russia, Vladimir Kolossov #233- Il primo quinquennio della repubblica di dicembre in Russia: come si costruiscono gli istituti e si strutturano gli interessi, Petr Fedossov #257- Lo stato attuale e le prospettive della Chiesa ortodossa in Russia, P. Innokentij Pavlov #272- La nuova influenza dei valori religiosi sull’intellighenzia russa (tendenze attuali), Andrei B. Zubov #295- La religiosità postsovietica: dall’eclettismo religioso alle fedi nazionali, Sergei Filatov #32

A case of structure determination using pseudosymmetry

When properly applied, pseudosymmetry can be used to improve crystallographic phases through averaging and to facilitate crystal structure determination

Structure of the ArsA ATPase: the catalytic subunit of a heavy metal resistance pump

Active extrusion is a common mechanism underlying detoxification of heavy metals, drugs and antibiotics in bacteria, protozoa and mammals. In Escherichia coli, the ArsAB pump provides resistance to arsenite and antimonite. This pump consists of a soluble ATPase (ArsA) and a membrane channel (ArsB). ArsA contains two nucleotide-binding sites (NBSs) and a binding site for arsenic or antimony. Binding of metalloids stimulates ATPase activity. The crystal structure of ArsA reveals that both NBSs and the metal-binding site are located at the interface between two homologous domains. A short stretch of residues connecting the metal-binding site to the NBSs provides a signal transduction pathway that conveys information on metal occupancy to the ATP hydrolysis sites. Based on these structural features, we propose that the metal-binding site is involved directly in the process of vectorial translocation of arsenite or antimonite across the membrane. The relative positions of the NBS and the inferred mechanism of allosteric activation of ArsA provide a useful model for the interaction of the catalytic domains in other transport ATPases

Structure and mechanism of 3-deoxy-d-manno-octulosonate 8-phosphate synthase

3-Deoxy-d-manno-octulosonate 8-phosphate (KDO8P) synthase catalyzes the condensation of phosphoenolpyruvate (PEP) with arabinose 5-phosphate (A5P) to form KDO8P and inorganic phosphate. KDO8P is the phosphorylated precursor of 3-deoxy-d-manno-octulosonate, an essential sugar of the lipopolysaccharide of Gram-negative bacteria. The crystal structure of the Escherichia coli KDO8P synthase has been determined by multiple wavelength anomalous diffraction and the model has been refined to 2.4 Å (R-factor, 19.9%; R-free, 23.9%). KDO8P synthase is a homotetramer in which each monomer has the fold of a (β/α)₈ barrel. On the basis of the features of the active site, PEP and A5P are predicted to bind with their phosphate moieties 13 Å apart such that KDO8P synthesis would proceed via a linear intermediate. A reaction similar to KDO8P synthesis, the condensation of phosphoenolpyruvate, and erythrose 4-phosphate to form 3-deoxy-d-arabino-heptulosonate 7-phosphate (DAH7P), is catalyzed by DAH7P synthase. In the active site of DAH7P synthase the two substrates PEP and erythrose 4-phosphate appear to bind in a configuration similar to that proposed for PEP and A5P in the active site of KDO8P synthase. This observation suggests that KDO8P synthase and DAH7P synthase evolved from a common ancestor and that they adopt the same catalytic strategy