A Backup Plan for Self-Protection: S-Methylation of Holomycin Biosynthetic Intermediates in Streptomyces clavuligerus

Biosynthesis of the dithiolopyrrolone antibiotic holomycin in Streptomyces clavuligerus involves the closure of a pair of enethiols to a cyclic disulfide. We have shown that the dithiol oxidase HlmI is responsible for the disulfide formation and this enzyme also plays a role in self-protection. In the present study, we examine how S. clavuligerus deals with the proposed toxic dithiol intermediates when hlmI is deleted. We used differential NMR spectroscopy and mass spectrometry to profile the metabolomes of hlmI deletion mutants along with the wild-type strain and a holomycin-overproducing strain. A number of metabolites unique to ΔhlmI strains were identified. In these metabolites the enethiols have been incapacitated by a combination of mono- and di-S-methylation. We also observed an intriguing dimeric thioether adduct in low quantities in the wild-type strain and at much higher levels in the ΔhlmI strains. The structures of these novel metabolites highlight the reactivity of the dihydrodithiolopyrrolone scaffold. Furthermore, bioassays suggest that modification of the enethiol warhead by S-alkylation provides a host strategy for detoxification, one that is shared amongst multiple species producing such bioactive disulfide natural products

Plasma exchange for primary autoimmune autonomic failure

We report on a patient with long-standing severe autonomic failure that affected his sympathetic and parasympathetic nervous systems. Antibodies against the ganglionic acetylcholine receptors were detected in the serum. Removal of the antibodies by means of plasma exchange resulted in a dramatic clinical improvement

Interception of the Bycroft-Gowland Intermediate in the Enzymatic Macrocyclization of Thiopeptides

Thiopeptides are a broad class of macrocyclic, heavily modified peptide natural products that are unified by the presence of a substituted, nitrogen-containing heterocycle core. Early work indicated that this core might be fashioned from two dehydroalanines by an enzyme-catalyzed aza-[4 + 2] cycloaddition to give a cyclic-hemiaminal intermediate. This common intermediate could then follow a reductive path toward a dehydropiperidine, as in the thiopeptide thiostrepton, or an aromatization path to yield the pyridine groups observed in many other thiopeptides. Although several of the enzymes proposed to perform this cycloaddition have been reconstituted, only pyridine products have been isolated and any hemiaminal intermediates have yet to be observed. Here, we identify the conditions and substrates that decouple the cycloaddition from subsequent steps and allow interception and characterization of this long hypothesized intermediate. Transition state modeling indicates that the key amide-iminol tautomerization is the major hurdle in an otherwise energetically favorable cycloaddition. An anionic model suggests that deprotonation and polarization of this amide bond by TbtD removes this barrier and provides a sufficient driving force for facile (stepwise) cycloaddition. This work provides evidence for a mechanistic link between disparate cyclases in thiopeptide biosynthesis

Phevamine A, a small molecule that suppresses plant immune responses

Bacterial plant pathogens cause significant crop damage worldwide. They invade plant cells by producing a variety of virulence factors, including small-molecule toxins and phytohormone mimics. Virulence of the model pathogen Pseudomonas syringae pv. tomato DC3000 (Pto) is regulated in part by the sigma factor HrpL. Our study of the HrpL regulon identified an uncharacterized, three-gene operon in Pto that is controlled by HrpL and related to the Erwinia hrp-associated systemic virulence (hsv) operon. Here, we demonstrate that the hsv operon contributes to the virulence of Pto on Arabidopsis thaliana and suppresses bacteria-induced immune responses. We show that the hsv-encoded enzymes in Pto synthesize a small molecule, phevamine A. This molecule consists of L-phenylalanine, L-valine, and a modified spermidine, and is different from known small molecules produced by phytopathogens. We show that phevamine A suppresses a potentiation effect of spermidine and L-arginine on the reactive oxygen species burst generated upon recognition of bacterial flagellin. The hsv operon is found in the genomes of divergent bacterial genera, including ∼37% of P. syringae genomes, suggesting that phevamine A is a widely distributed virulence factor in phytopathogens. Our work identifies a small-molecule virulence factor and reveals a mechanism by which bacterial pathogens overcome plant defense. This work highlights the power of omics approaches in identifying important small molecules in bacteria-host interactions

Time-integrated luminosity recorded by the BABAR detector at the PEP-II e+e- collider

This article is the Preprint version of the final published artcile which can be accessed at the link below.We describe a measurement of the time-integrated luminosity of the data collected by the BABAR experiment at the PEP-II asymmetric-energy e+e- collider at the ϒ(4S), ϒ(3S), and ϒ(2S) resonances and in a continuum region below each resonance. We measure the time-integrated luminosity by counting e+e-→e+e- and (for the ϒ(4S) only) e+e-→μ+μ- candidate events, allowing additional photons in the final state. We use data-corrected simulation to determine the cross-sections and reconstruction efficiencies for these processes, as well as the major backgrounds. Due to the large cross-sections of e+e-→e+e- and e+e-→μ+μ-, the statistical uncertainties of the measurement are substantially smaller than the systematic uncertainties. The dominant systematic uncertainties are due to observed differences between data and simulation, as well as uncertainties on the cross-sections. For data collected on the ϒ(3S) and ϒ(2S) resonances, an additional uncertainty arises due to ϒ→e+e-X background. For data collected off the ϒ resonances, we estimate an additional uncertainty due to time dependent efficiency variations, which can affect the short off-resonance runs. The relative uncertainties on the luminosities of the on-resonance (off-resonance) samples are 0.43% (0.43%) for the ϒ(4S), 0.58% (0.72%) for the ϒ(3S), and 0.68% (0.88%) for the ϒ(2S).This work is supported by the US Department of Energy and National Science Foundation, the Natural Sciences and Engineering Research Council (Canada), the Commissariat à l’Energie Atomique and Institut National de Physique Nucléaire et de Physiquedes Particules (France), the Bundesministerium für Bildung und Forschung and Deutsche Forschungsgemeinschaft (Germany), the Istituto Nazionale di Fisica Nucleare (Italy), the Foundation for Fundamental Research on Matter (The Netherlands), the Research Council of Norway, the Ministry of Education and Science of the Russian Federation, Ministerio de Ciencia e Innovación (Spain), and the Science and Technology Facilities Council (United Kingdom). Individuals have received support from the Marie-Curie IEF program (European Union) and the A.P. Sloan Foundation (USA)

The <i>Ectocarpus</i> genome and the independent evolution of multicellularity in brown algae

Brown algae (Phaeophyceae) are complex photosynthetic organisms with a very different evolutionary history to green plants, to which they are only distantly related1. These seaweeds are the dominant species in rocky coastal ecosystems and they exhibit many interesting adaptations to these, often harsh, environments. Brown algae are also one of only a small number of eukaryotic lineages that have evolved complex multicellularity (Fig. 1).We report the 214 million base pair (Mbp) genome sequence of the filamentous seaweed Ectocarpus siliculosus (Dillwyn) Lyngbye, a model organism for brown algae, closely related to the kelps (Fig. 1). Genome features such as the presence of an extended set of light-harvesting and pigment biosynthesis genes and new metabolic processes such as halide metabolism help explain the ability of this organism to cope with the highly variable tidal environment. The evolution of multicellularity in this lineage is correlated with the presence of a rich array of signal transduction genes. Of particular interest is the presence of a family of receptor kinases, as the independent evolution of related molecules has been linked with the emergence of multicellularity in both the animal and green plant lineages. The Ectocarpus genome sequence represents an important step towards developing this organism as a model species, providing the possibility to combine genomic and genetic2 approaches to explore these and other aspects of brown algal biology further

Observation of the baryonic decay B \uaf 0 \u2192 \u39bc+ p \uaf K-K+

We report the observation of the baryonic decay B\uaf0\u2192\u39bc+p\uafK-K+ using a data sample of 471 7106 BB\uaf pairs produced in e+e- annihilations at s=10.58GeV. This data sample was recorded with the BABAR detector at the PEP-II storage ring at SLAC. We find B(B\uaf0\u2192\u39bc+p\uafK-K+)=(2.5\ub10.4(stat)\ub10.2(syst)\ub10.6B(\u39bc+)) 710-5, where the uncertainties are statistical, systematic, and due to the uncertainty of the \u39bc+\u2192pK-\u3c0+ branching fraction, respectively. The result has a significance corresponding to 5.0 standard deviations, including all uncertainties. For the resonant decay B\uaf0\u2192\u39bc+p\uaf\u3c6, we determine the upper limit B(B\uaf0\u2192\u39bc+p\uaf\u3c6)<1.2 710-5 at 90% confidence level

Search for Darkonium in e+e- Collisions

Collider searches for dark sectors, new particles interacting only feebly with ordinary matter, have largely focused on identifying signatures of new mediators, leaving much of dark sector structures unexplored. In particular, the existence of dark matter bound states (darkonia) remains to be investigated. This possibility could arise in a simple model in which a dark photon (A0 ) is light enough to generate an attractive force between dark fermions. We report herein a search for a JPC ¼ 1−− darkonium state, the ϒD, produced in the reaction eþe− → γϒD, ϒD → A0 A0 A0 , where the dark photons subsequently decay into pairs of leptons or pions, using 514 fb−1 of data collected with the BABAR detector. No significant signal is observed, and we set bounds on the γ − A0 kinetic mixing as a function of the dark sector coupling constant for 0.001 < mA0 < 3.16 GeV and 0.05 < mϒD < 9.5 GeV.publishedVersio

Operation and performance of the ATLAS semiconductor tracker

The semiconductor tracker is a silicon microstrip detector forming part of the inner tracking system of the ATLAS experiment at the LHC. The operation and performance of the semiconductor tracker during the first years of LHC running are described. More than 99% of the detector modules were operational during this period, with an average intrinsic hit efficiency of (99.74±0.04)%. The evolution of the noise occupancy is discussed, and measurements of the Lorentz angle, δ-ray production and energy loss presented. The alignment of the detector is found to be stable at the few-micron level over long periods of time. Radiation damage measurements, which include the evolution of detector leakage currents, are found to be consistent with predictions and are used in the verification of radiation background simulations