Expression, purification, crystallization and preliminary X-ray diffraction analysis of conjugated bile salt hydrolase from Bifidobacterium longum

Conjugated bile salt hydrolase (BSH) catalyses the hydrolysis of the amide bond that conjugates bile acids to glycine and to taurine. The BSH enzyme from Bifidobacterium longum was overexpressed in Escherichia coli BL21(DE3), purified and crystallized. Crystallization conditions were screened using the hanging-drop vapour-diffusion method. Crystal growth, with two distinct morphologies, was optimal in experiments carried out at 303 K. The crystals belong to the hexagonal system, space group P622 with unit-cell parameters a = b = 124.86, c = 219.03 Angstrom, and the trigonal space group P321, with unit-cell parameters a = b = 125.24, c = 117.03 Angstrom. The crystals diffracted X-rays to 2.5 Angstrom spacing. Structure determination using the multiple isomorphous replacement method is in progress

Expression, purification, crystallization and preliminary X-ray diffraction analysis of conjugated bile salt hydrolase from Bifidobacterium longum

Conjugated bile salt hydrolase (BSH) catalyses the hydrolysis of the amide bond that conjugates bile acids to glycine and to taurine. The BSH enzyme from Bifidobacterium longum was overexpressed in Escherichia coli BL21(DE3), purified and crystallized. Crystallization conditions were screened using the hanging-drop vapour-diffusion method. Crystal growth, with two distinct morphologies, was optimal in experiments carried out at 303 K. The crystals belong to the hexagonal system, space group P622 with unit-cell parameters a = b = 124.86, c = 219.03 Angstrom, and the trigonal space group P321, with unit-cell parameters a = b = 125.24, c = 117.03 Angstrom. The crystals diffracted X-rays to 2.5 Angstrom spacing. Structure determination using the multiple isomorphous replacement method is in progress

X-ray Spectrum and Pulsations of the Vela Pulsar

We report the results of the spectral and timing analysis of observations of the Vela pulsar with the Chandra X-ray Observatory. The spectrum shows no statistically significant spectral lines in the observed 0.25--8.0 keV band. It consists of two distinct continuum components. The softer component can be modeled as either a magnetic hydrogen atmosphere spectrum with kT = 59 +- 3 eV, R = 15.5 +- 1.5 km, or a standard blackbody with kT = 129 +- 4 eV, R = 2.1 +- 0.2 km (the radii are for a distance of 250 pc). The harder component, modeled as a power-law spectrum, gives photon indices depending on the model adopted for the soft component: gamma = 1.5 +- 0.3 for the magnetic atmosphere soft component, and gamma = 2.7 +- 0.4 for the blackbody soft component. Timing analysis shows three peaks in the pulse profile, separated by about 0.3 in phase. Energy-resolved timing provides evidence for pulse profile variation with energy. The higher energy (E > 1.8 keV) profile shows significantly higher pulsed fraction.Comment: 4 pages, 2 figures, To appear in "Neutron Stars in Supernova Remnants" (ASP Conference Proceedings), eds P. O. Slane and B. M. Gaensler Corrected TYPO

Astrometry of OH/IR stars using 1612 MHz hydroxyl masers. I. Annual parallaxes of WX Psc and OH138.0+7.2

We report on the measurement of the trigonometric parallaxes of 1612 MHz hydroxyl masers around two asymptotic giant branch stars, WX Psc and OH138.0+7.2, using the NRAO Very Long Baseline Array with in-beam phase referencing calibration. We obtained a 3-sigma upper limit of <=5.3 mas on the parallax of WX Psc, corresponding to a lower limit distance estimate of >~190 pc. The obtained parallax of OH138.0+7.2 is 0.52+/-0.09 mas (+/-18%), corresponding to a distance of 1.9(+0.4,-0.3) kpc, making this the first hydroxyl maser parallax below one milliarcsecond. We also introduce a new method of error analysis for detecting systematic errors in the astrometry. Finally, we compare our trigonometric distances to published phase-lag distances toward these stars and find a good agreement between the two methods.Comment: Preprint, accepted for publication in The Astronomical Journal (January 17, 2017

Correlation between X-ray Lightcurve Shape and Radio Arrival Time in the Vela Pulsar

We report the results of simultaneous observations of the Vela pulsar in X-rays and radio from the RXTE satellite and the Mount Pleasant Radio Observatory in Tasmania. We sought correlations between the Vela's X-ray emission and radio arrival times on a pulse by pulse basis. At a confidence level of 99.8% we have found significantly higher flux density in Vela's main X-ray peak during radio pulses that arrived early. This excess flux shifts to the 'trough' following the 2nd X-ray peak during radio pulses that arrive later. Our results suggest that the mechanism producing the radio pulses is intimately connected to the mechanism producing X-rays. Current models using resonant absorption of radio emission in the outer magnetosphere as a cause of the X-ray emission are explored as a possible explanation for the correlation.Comment: 6 pages, 5 figures, accepted by Ap

The structures of Micrococcus lysodeikticus catalase, its ferryl intermediate (compound II) and NADPH complex

The crystal structure of the bacterial catalase from Micrococcus lysodeikticus has been refined using the gene-derived sequence both at 0.88 Angstrom resolution using data recorded at 110 K and at 1.5 Angstrom resolution with room-temperature data. The atomic resolution structure has been refined with individual anisotropic atomic thermal parameters. This has revealed the geometry of the haem and surrounding protein, including many of the H atoms, with unprecedented accuracy and has characterized functionally important hydrogen-bond interactions in the active site. The positions of the H atoms are consistent with the enzymatic mechanism previously suggested for beef liver catalase. The structure reveals that a 25 Angstrom long channel leading to the haem is filled by partially occupied water molecules, suggesting an inherent facile access to the active site. In addition, the structures of the ferryl intermediate of the catalase, the so-called compound II, at 1.96 Angstrom resolution and the catalase complex with NADPH at 1.83 Angstrom resolution have been determined. Comparison of compound II and the resting state of the enzyme shows that the binding of the O atom to the iron (bond length 1.87 Angstrom) is associated with increased haem bending and is accompanied by a distal movement of the iron and the side chain of the proximal tyrosine. Finally, the structure of the NADPH complex shows that the cofactor is bound to the molecule in an equivalent position to that found in beef liver catalase, but that only the adenine part of NADPH is visible in the present structure