Neutralization of the anthrax toxin by antibody-mediated stapling of its membrane-penetrating loop.

Anthrax infection is associated with severe illness and high mortality. Protective antigen (PA) is the central component of the anthrax toxin, which is one of two major virulence factors of Bacillus anthracis, the causative agent of anthrax disease. Upon endocytosis, PA opens a pore in the membranes of endosomes, through which the cytotoxic enzymes of the toxin are extruded. The PA pore is formed by a cooperative conformational change in which the membrane-penetrating loops of PA associate, forming a hydrophobic rim that pierces the membrane. Due to its crucial role in anthrax progression, PA is an important target for monoclonal antibody-based therapy. cAb29 is a highly effective neutralizing antibody against PA. Here, the cryo-EM structure of PA in complex with the Fab portion of cAb29 was determined. It was found that cAb29 neutralizes the toxin by clamping the membrane-penetrating loop of PA to the static surface-exposed loop of the D3 domain of the same subunit, thereby preventing pore formation. These results provide the structural basis for the antibody-based neutralization of PA and bring into focus the membrane-penetrating loop of PA as a target for the development of better anti-anthrax vaccines.

Achilles tendon shape and echogenicity on ultrasound among active badminton players.

The relationship between Achilles tendon ultrasound abnormalities, including a spindle shape and heterogeneous echogenicity, is unclear. This study investigated the relationship between these abnormalities, tendon thickness, Doppler flow and pain. Sixty-one badminton players (122 tendons, 36 men, and 25 women) were recruited. Achilles tendon thickness, shape (spindle, parallel), echogenicity (heterogeneous, homogeneous) and Doppler flow (present or absent) were measured bilaterally with ultrasound. Achilles tendon pain (during or after activity over the last week) and pain and function [Victorian Institute of Sport Achilles Assessment (VISA-A)] were measured. Sixty-eight (56%) tendons were parallel with homogeneous echogenicity (normal), 22 (18%) were spindle shaped with homogeneous echogenicity, 16 (13%) were parallel with heterogeneous echogenicity and 16 (13%) were spindle shaped with heterogeneous echogenicity. Spindle shape was associated with self-reported pain (P<0.05). Heterogeneous echogenicity was associated with lower VISA-A scores than normal tendon (P<0.05). There was an ordinal relationship between normal tendon, parallel and heterogeneous and spindle shaped and heterogeneous tendons with regard to increasing thickness and likelihood of Doppler flow. Heterogeneous echogenicity with a parallel shape may be a physiological phase and may develop into heterogeneous echogenicity with a spindle shape that is more likely to be pathological.

Structural insights into the design of inhibitors for the L1 metallo-beta-lactamase from Stenotrophomonas maltophilia.

One mechanism by which bacteria can escape the action of beta-lactam antibiotics is the production of metallo-beta-lactamases. Inhibition of these enzymes should restore the action of these widely used antibiotics. The tetrameric enzyme L1 from Stenotrophomonas maltophilia was used as a model system to determine a series of high-resolution crystal structures of apo, mono and bi-metal substituted proteins as well as protein-inhibitor complexes. Unexpectedly, although the apo structure revealed only few significant structural differences from the holo structure, some inhibitors were shown to induce amino acid side-chain rotations in the tightly packed active site. Moreover, one inhibitor employs a new binding mode in order to interact with the di-zinc center. This structural information could prove essential in the process of elucidation of the mode of interaction between a putative lead compound and metallo-beta-lactamases, one of the main steps in structure-based drug design.

Protonation state and substrate binding to B2 metallo-beta-lactamase CphA from Aeromonas hydrofila.

The zinc enzymes metallo beta-lactamases counteract the beneficial action of beta-lactam antibiotics against bacterial infections, by hydrolyzing their beta-lactam rings. To understand structure/function relationships on a representative member of this class, the B2 M beta L CphA, we have investigated the H-bond pattern at the Zn enzymatic active site and substrate binding mode by molecular simulation methods. Extensive QM calculations at the DFT-BLYP level on eleven models of the protein active site, along with MD simulations of the protein in aqueous solution, allow us to propose two plausible protonation states for the unbound enzyme, which are probably in equilibrium. Docking procedures along with MD simulations and QM calculations suggest that in the complex between the enzyme and its substrate (biapenem), the latter is stable in only one of the two protonation states, in addition it exhibits two different binding modes, of which only one agrees with previous proposals. In both cases, the substrate is polarized as in aqueous solution. We conclude that addressing mechanistic issues on this class of enzymes requires a careful procedure to assign protonation states and substrate docking modes.

Competitive inhibitors of the CphA metallo-beta-lactamase from Aeromonas hydrophila.

Various inhibitors of metallo-beta-lactamases have been reported; however, none are effective for all subgroups. Those that have been found to inhibit the enzymes of subclass B2 (catalytically active with one zinc) either contain a thiol (and show less inhibition towards this subgroup than towards the dizinc members of B1 and B3) or are inactivators behaving as substrates for the dizinc family members. The present work reveals that certain pyridine carboxylates are competitive inhibitors of CphA, a subclass B2 enzyme. X-ray crystallographic analyses demonstrate that pyridine-2,4-dicarboxylic acid chelates the zinc ion in a bidentate manner within the active site. Salts of these compounds are already available and undergoing biomedical testing for various nonrelated purposes. Pyridine carboxylates appear to be useful templates for the development of more-complex, selective, nontoxic inhibitors of subclass B2 metallo-beta-lactamases.

Crystallization and preliminary X-ray diffraction analysis of human transcobalamin, a vitamin B12-transporting protein.

Transcobalamin is a cobalamin-binding protein in mammalian plasma that facilitates the cellular uptake of vitamin B(12). Human transcobalamin was crystallized using polyethylene glycol and ethanol as precipitants. Crystals belong to the orthorhombic space group P2(1)2(1)2(1), with unit-cell parameters a = 49.04, b = 145.27, c = 164.96 A. A complete data set to 3.2 A resolution was collected from a single crystal using synchrotron radiation. Estimation of the crystal packing (V(M) = 3.2 A(3) Da(-1)) and self-rotation function analysis suggest the presence of two molecules in the asymmetric unit related by non-crystallographic twofold symmetry.

Crystallization and preliminary X-ray analysis of two pH-dependent forms of cytochrome c2 from Rhodopseudomonas palustris.

Cytochrome c(2) from Rhodopseudomonas palustris has been crystallized in two different crystal forms: a monoclinic form I at pH 4.4 from both reduced and oxidized protein solution and a trigonal form II at pH 9.0 from reduced protein solution. Complete 1. 7 and 1.4 A resolution data sets were collected from the oxidized form I and from the form II, respectively. The preliminary structures show an important change in the iron coordination environment in the trigonal form obtained at basic pH arising from the substitution of the Met ligand by an ammonia molecule.

[Urinary infection in patients with short-term bladder catheterization]. / Estudio de la infección urinaria en pacientes con sondaje vesical de corta duración.

BACKGROUND: The aim of the study was to assess the number of urinary tract infections, the time between catheterization and the development of bacteriuria, the route taken by the microorganisms and the clinical relevance of infection in patients with bladder catheterization during a short time. METHODS: 83 patients were evaluated by urine culture from the bladder, urethra, the connection of the catheter and the collecting system, and the collecting bag, on a daily basis during the 5 initial days, with clinical and microbiological follow up to 7 days after the removal of the catheter. RESULTS: 32% of patients developed bacteriuria. The mean interval between catheterization and the development of bacteriuria was 4.1 days. 45% of them were detected within 48 hours after catheter insertion. In 50% the causative organisms were detected in the urethra at least 24 hours previously. In no case the microorganism isolated from the connection between the catheter and the collecting system was subsequently isolated form bladder urine. Bacteriuria disappeared spontaneously in 27% of cases. Antibiotic therapy was necessary in 42%. CONCLUSIONS: In the study population most urinary infections were due to previous urethral colonization and subsequent dragging into the bladder, in many cases at the same time of catheterization. A non-negligible number of bacteriurias disappeared spontaneously.

Efficacy of teicoplanin and imipenem on respiratory infections in high risk patients.

The efficacy of teicoplanin and imipenem was assayed on microorganisms, mainly hospital opportunistic organisms, resistant to common therapy, isolated from bronchoaspirates of high risk hospitalized patients. Imipenem has shown remarkable activity against all isolated microorganisms with sensitivities ranging from 91.93% against Acinetobacter calcoaceticus to 75.86% against Pseudomonas sp. Teicoplanin has shown excellent activity against Staphylococcus aureus (sensitivity 83.66%).

[Autochthonous mycetoma caused by Petriellidium boydii (author's transl)]. / Micetoma autóctono por Petriellidium boydii.

A case of authochthonous mycetoma with Petriellidium boydii as the causal agent, located in the third distal of the lower left leg is reported. The patient was a 35 year old male without any associated pathology. It is assumed that the infecting organism entered the tissues through a lesion caused a few years previously during a traffic accident. On admittance patient was found to have two fistulas on the upper part of the external malleolus from which pus with small yellow granules measuring 1 to 1.5 mm in diameter discharged. Fungus was repeatedly isolated in pure culture in all media in which is was sown. Mycological study of the isolated fungus and histopathology of the lesion are presented. The fungus proved in vitro to be sensitive to miconazole nitrate with minimal inhibitory concentration and minimal fungicide concentration values of less than 0.1 microgram/ml. The third distal of the peroneal bone was excised and arthrodesis of the ankle practiced as conservative treatment and miconazole administered intravenously, Eighteen months later clinical evolution is satisfactory.

[Anicteric calculosis of the common bile duct]. / La calcolosi anitterica dell'epato-coledoco.

In a series of 158 patients who underwent surgical treatment for biliary lithiasis, we have had 109 (69%) lithiasis of the gallbladder (CC) and 48 (31%) lithiasis of the common bile duct. In the last group 19 (38%) were without jaundice (CAEC) and 30 (62%) with jaundice (CIEC). We have compared the three group of patients (CC, CAEC and CIEC). It has come out that CAEC is between CC and CIEC and that attacks of biliary fever and high levels of alkaline phosphatase and transaminases in the serum are the helpful findings for preoperative diagnosis. Intravenous cholangiograms can confirm the diagnosis in 80% of one series but the operative cholangiography remains the definitive test. Finally CAEC seems to be the "benign" lithiasis of the common bile duct, a first stage that getting worse will became CIEC.