Cell Surface Labeling of Escherichia coli via Copper(I)-Catalyzed [3+2] Cycloaddition

Labeling of the cell surface of Escherichia coli was accomplished by expression of a recombinant outer membrane protein, OmpC, in the presence of the unnatural amino acid azidohomoalanine, which acts as a methionine surrogate. The surface-exposed azide moieties of whole cells were biotinylated via Cu(1)-catalyzed [3+2] azide-alkyne cycloaddition. The specificity of labeling of both wild-type OmpC and a mutant containing additional methionine sites for azidohomoalanine incorporation was confirmed by Western blotting. Flow cytometry was performed to examine the specificity of the labeling. Cells that express the mutant form of OmpC in the presence of azidohomoalanine, which were biotinylated and stained with fluorescent avidin, exhibit a mean fluorescence 10-fold higher than the background. Incorporation of an unnatural amino acid can thus be determined on a single-cell basis

Introduction of an Aliphatic Ketone into Recombinant Proteins in a Bacterial Strain that Overexpresses an Editing-Impaired Leucyl-tRNA Synthetase

Like leucine? A leucine analogue containing a ketone has been incorporated into proteins in E. coli. Only E. coli strains overexpressing an editing-deficient leucyl-tRNA synthetase were capable of synthesizing proteins with the aliphatic ketone amino acid. Modification of ketone-containing proteins under mild conditions has been demonstrated

Pulsar Constraints on Neutron Star Structure and Equation of State

With the aim of constraining the structural properties of neutron stars and the equation of state of dense matter, we study sudden spin-ups, glitches, occurring in the Vela pulsar and in six other pulsars. We present evidence that glitches represent a self-regulating instability for which the star prepares over a waiting time. The angular momentum requirements of glitches in Vela indicate that at least 1.4% of the star's moment of inertia drives these events. If glitches originate in the liquid of the inner crust, Vela's `radiation radius' must exceed ~12 km for a mass of 1.4 solar masses. Observational tests of whether other neutron stars obey this constraint will be possible in the near future.Comment: 5 pages, including figures. To appear in Physical Review Letter

Outcomes for patients with severe chronic neutropenia treated with granulocyte colony-stimulating factor

Severe chronic neutropenia (SCN), defined as blood neutrophils \u3c0.5 × 109/L for \u3e3 months, is an uncommon hematological condition associated with recurrent and severe bacterial infections. After short-term clinical trials showed the benefits of granulocyte colony-stimulating factor (G-CSF) treatment for SCN, SCNIR (Severe Chronic Neutropenia International Registry) opened to determine the long-term benefits and safety of this treatment. This report summarizes findings from more than 16 000 patient-years of prospective observations for patients with congenital and acquired SCN. We observed that adverse outcomes depend on the underlying etiology. Myelodysplasia (MDS) and acute myeloid leukemia (AML) occur infrequently and largely in patients with congenital neutropenias. Having cyclic or chronic autoimmune/ idiopathic neutropenia portends a favorable prognosis. A few patients with idiopathic neutropenia evolve to develop lymphoid malignancies, but they do not appear to be at increased risk of myeloid malignancies, even with very long-term G-CSF therapy. Progression to systemic autoimmune diseases, bone marrow (BM) failure, aplastic anemia, or nonmyeloid malignancies are not expected consequences of SCN or treatment with G-CSF

Simulated Performance of 3-DTI Gamma-Ray Telescope Concepts

We present Monte Carlo simulations of two astronomical gamma-ray telescope concepts based on the ThreeDimensional Track Imager (3- DTI) detector. The 3-DTI consists of a time projection chamber with two-dimensional, crossedstrip micro-well detector readout. The full three- dimensional reconstruction of charged-particle tracks in the gas volume is obtained from transient digitizers, which record the time signature of the charge collected in the wells of each strip. Such detectors hold great promise for advanced Compton telescope (ACT) and advanced pair telescope (APT) concepts due to the very precise measurement of charged particle momenta that is possible (Compton recoil electrons and electron-positron pairs, respectively). We have investigated the performance of baseline ACT and APT designs based on the 3-DTI detector using simulation tools based on GEANT3 and GEANT4, respectively. We present the expected imaging, spectroscopy, polarimetry, and background performance of each design

Medium-Energy Gamma-Ray Astrophysics with the 3-DTI Gamma-Ray Telescope

Gamma-ray observations in the medium energy range (0.50-50.0 MeV) are central to unfolding many outstanding questions in astrophysics. The challenges of medium-energy gamma-ray observations, however, are the low photon statistics and large backgrounds. We review these questions, address the telescope technology requirements, and describe our development of the 3-Dimensional Track Imaging (3-DTI) Compton telescope and its performance for a new mediumenergy gamma-ray mission. The 3-DTI is a large-volume time projection chamber (TPC) with a 2-dimensional gas micro-well detector (MWD) readout

Lamotrigine Overdose Presenting as Shock and Pulmonary Edema

Abstract: Lamotrigine is a broad-spectrum anti-epileptic drug with a good safety profile used to treat general and focal epilepsy. Toxicity is uncommon and usually mild, and symptoms usually include rash, headache, nausea, abdominal pain, somnolence, dizziness, and aggravated seizure activity. More severe adverse reactions are rare, but have been reported and include encephalopathy, hypotension, wide complex tachycardia, cardiac arrest, and death. Lamotrigine drug levels do not consistently correlate with either therapeutic effect or toxicity, though higher levels are thought to have a higher risk of toxicity. Also, the level is typically a send out test with a 1 to 3 day turnaround time. This case report details a case of Lamotrigine toxicity in a thirteen-year-old Hispanic female with moderately well-controlled focal epilepsy presenting with acute florid pulmonary edema and fluid refractory vasodilatory shock. These symptoms have not been reported together elsewhere in the pediatric literature. By expanding upon the known presentation of Lamotrigine toxicity in children, the time to diagnosis for future cases may be shortened, providers may avoid anchoring bias, and morbidity may decrease

Exploring Halo Substructure with Giant Stars. VI. Extended Distributions of Giant Stars Around the Carina Dwarf Spheroidal Galaxy -- How Reliable Are They?

The question of the existence of active tidal disruption around various dSph galaxies remains controversial. That debate often centers on the nature (bound vs. unbound) of extended populations of stars. However, the more fundamental issue of the very existence of the extended populations is still contentious. We present an evaluation of the debate centering on one particular dSph, Carina, for which claims both for and against the existence of stars beyond the King radius have been made. Our review includes an examination of all previous studies bearing on the Carina radial profile and shows that the survey method which achieves the highest detected dSph signal-to-background in the outer parts of the galaxy is the Washington M, T2 + DDO51 (MTD) filter approach from Paper II in this series. We then address statistical methods used to evaluate the reliability of MTD surveys in the presence of photometric errors and for which a new, a posteriori statistical analysis methodology is provided. Finally, these statistical methods are tested by new spectroscopy of stars in the MTD-selected Carina candidate sample. Of 74 candidate giants with follow-up spectroscopy, the MTD technique identified 61 new Carina members, including 8 stars outside the King radius. From a sample of 29 stars not initially identified as candidate Carina giants but that lie just outside of our selection criteria, 12 have radial velocities consistent with membership, including 5 extratidal stars. Carina is shown to have an extended population of giant stars extending to a major axis radius of 40' (1.44x the nominal King radius).Comment: 56 pages, 10 figures. Submitted to the Astronomical Journal, 2004 Sep 2

Unnatural Amino Acid Incorporation into Virus-Like Particles

Virus-like particles composed of hepatitis B virus (HBV) or bacteriophage Qβ capsid proteins have been labeled with azide- or alkyne-containing unnatural amino acids by expression in a methionine auxotrophic strain of E. coli. The substitution does not affect the ability of the particles to self-assemble into icosahedral structures indistinguishable from native forms. The azide and alkyne groups were addressed by Cu(I)-catalyzed [3 + 2] cycloaddition: HBV particles were decomposed by the formation of more than 120 triazole linkages per capsid in a location-dependent manner, whereas Qβ suffered no such instability. The marriage of these well-known techniques of sense-codon reassignment and bioorthogonal chemical coupling provides the capability to construct polyvalent particles displaying a wide variety of functional groups with near-perfect control of spacing