Assembly line enzymology by multimodular nonribosomal peptide synthetases: the thioesterase domain of E. coli EntF catalyzes both elongation and cyclolactonization

BackgroundEntF is a 142 kDa four domain (condensation-adenylationpeptidyl carrier protein-thioesterase) nonribosomal peptide synthetase (NRPS) enzyme that assembles the Escherichia coli N-acyl-serine trilactone siderophore enterobactin from serine, dihydroxybenzoate (DHB) and ATP with three other enzymes (EntB, EntD and EntE). To assess how EntF forms three ester linkages and cyclotrimerizes the covalent acyl enzyme DHB-Ser-S-PCP (peptidyl carrier protein) intermediate, we mutated residues of the proposed catalytic Ser-His-Asp triad of the thioesterase (TE) domain.ResultsThe Ser1138→Cys mutant (kcat decreased 1000-fold compared with wild-type EntF) releases both enterobactin (75%) and linear (DHB-Ser)2 dimer (25%) as products. The HiResultThe Ser1138→Cys mutant (kcat decreased 1000-fold compared with wild-type EntF) releases both enterobactin (75%) and linear (DHB-Ser)2 dimer (25%) as products. The His1271→Ala mutant (kcat decreased 10,000-fold compared with wild-type EntF) releases only enterobactin, but accumulates both DHB-Ser-O-TE and (DHB-Ser)2-O-TE acyl enzyme intermediates. Electrospray ionization and Fourier transform mass spectrometry of proteolytic digests were used to analyze the intermediates.71→Ala mutant (kcat decreased 10,000-fold compared with wild-type EntF) releases only enterobactin, but accumulates both DHB-Ser-O-TE and (DHB-Ser)2-O-TE acyl enzyme intermediates. Electrospray ionization and Fourier transform mass spectrometry of proteolytic digests were used to analyze the intermediates.ConclusionsThese results establish that the TE domain of EntF is both a cyclotrimerizing lactone synthetase and an elongation catalyst for ester-bond formation between covalently tethered DHB-Ser moieties, a new function for chain-termination TE domains found at the carboxyl termini of multimodular NRPSs and polyketide synthases

A web-based clinical decision tool to support treatment decision-making in psychiatry: a pilot focus group study with clinicians, patients and carers

Background. Treatment decision tools have been developed in many fields of medicine, including psychiatry, however benefits for patients have not been sustained once the support is withdrawn. We have developed a web-based computerised clinical decision support tool (CDST), which can provide patients and clinicians with continuous, up-to-date, personalised information about the efficacy and tolerability of competing interventions. To test the feasibility and acceptability of the CDST we conducted a focus group study, aimed to explore the views of clinicians, patients and carers. Methods. The CDST was developed in Oxford. To tailor treatments at an individual level, the CDST combines the best available evidence from the scientific literature with patient preferences and values, and with patient medical profile to generate personalised clinical recommendations. We conducted three focus groups comprising of three different participant types: consultant psychiatrists, participants with mental health diagnosis and/or experience of caring for someone with a mental health diagnosis, and primary care practitioners and nurses. Each 1-hour focus group started with a short visual demonstration of the CDST. To standardise the discussion during the focus groups, we used the same topic guide that covered themes relating to the acceptability and usability of the CDST. Focus groups were recorded and any identifying participant details were anonymised. Data were analysed thematically and managed using the Framework method and the constant comparative method. Results. The focus groups took place in Oxford between October 2016 and January 2017. Overall 31 participants attended (12 consultants, 11 primary care practitioners and 8 patients or carers). The main themes that emerged related to CDST applications in clinical practice, communication, conflicting priorities and record keeping. CDST was considered a useful clinical decision support, with recognised value in promoting clinician-patient collaboration and contributing to the development of personalised medicine. One major benefit of the CDST was perceived to be the open discussion about the possible side-effects of medications. Participants from all the three groups, however, universally commented that the terminology and language presented on the CDST were too medicalised, potentially leading to ethical issues around consent to treatment. Conclusions. The CDST can improve communication pathways between patients, carers and clinicians, identifying care priorities and providing an up-to-date platform for implementing evidence-based practice, with regard to prescribing practices

Case management and Think First completion

“The final, definitive version of this article has been published in the Journal, Probation Journal, Vol 53 Issue 3, 2006, Copyright The Trade Union and Professional Association for Family Court and Probation Staff, by SAGE Publications Ltd at: http://prb.sagepub.com/ " DOI: 10.1177/0264550506066771This article considers the findings of a small-scale study of the practice of case managers supervising offenders required to attend the Think First Group. It explores the interface between one-to-one and group-based work within multi-modal programmes of supervision and seeks to identify those practices that support individuals in completing a group.Peer reviewe

The use of interpretive phenomenological analysis in couple and family therapy research

This article proposes a research methodology that is newer to the field of couple and family therapy research called Interpretive Phenomenological Analysis (IPA). Researchers exploring couple and family therapy research continue to establish the efficacy of couple and family interventions in a context that favors a positivist view of phenomena. This research continues to be critical for establishing the role of couple and family therapy in the field of mental health as well as further clarifying which interventions are best for specific clinical issues and when. IPA offers researchers the opportunity to explore how couples and families make meaning of their experiences from an intersubjective perspective. Meaning making is central to understanding couples and families as well as part of the many clinical approaches to working with couples and families. Despite the importance of meaning, few research methodologies allow for this central concept in couple and family therapy to be the focus of exploration. The following article outlines one such methodology and the possible use of IPA in couple and family therapy research

Small-spatial scale variations of nebular properties and the abundance discrepancy in three Galactic HII regions

We present results of long-slit spectroscopy in several slit positions that cover different morphological structures of the central parts of three bright Galactic HII regions: M8, M17 and NGC7635. We study the spatial distributions of a large number of nebular parameters such as the extinction coefficient, line fluxes, physical conditions and ionic abundances at the maximum spatial resolution attainable with our instrumentation. Particularly, our goal is to study the behaviour of the abundance discrepancy factor of O^{2+}, ADF(O^{2+}), defined as the logarithmic difference of the O^{2+} abundances derived from collisionally excited and recombination lines. We find that the ADF(O^{2+}) remains fairly constant along the slit positions of M8 and M17. In the case of NGC7635, we only detect the OII recombination lines in the integrated spectrum along the whole slit, where the ADF(O^{2+}) reaches a remarkably high value of about 0.59 dex. We compare our results with previous ones obtained for the Orion Nebula. We find several evidences that suggest the presence of a candidate to Herbig-Haro object in M8.Comment: 18 pages, 9 figures. Table A1 available only in the online version. Accepted for publication in MNRAS

Channeling by Proximity: The Catalytic Advantages of Active Site Colocalization Using Brownian Dynamics

Nature often colocalizes successive steps in a metabolic pathway. Such organization is predicted to increase the effective concentration of pathway intermediates near their recipient active sites and to enhance catalytic efficiency. Here, the pathway of a two-step reaction is modeled using a simple spherical approximation for the enzymes and substrate particles. Brownian dynamics are used to simulate the trajectory of a substrate particle as it diffuses between the active site zones of two different enzyme spheres. The results approximate distances for the most effective reaction pathways, indicating that the most effective reaction pathway is one in which the active sites are closely aligned. However, when the active sites are too close, the ability of the substrate to react with the first enzyme was hindered, suggesting that even the most efficient orientations can be improved for a system that is allowed to rotate or change orientation to optimize the likelihood of reaction at both sites

The Atacama Cosmology Telescope: Cosmological Parameters from the 2008 Power Spectra

We present cosmological parameters derived from the angular power spectrum of the cosmic microwave background (CMB) radiation observed at 148 GHz and 218 GHz over 296 deg^2 with the Atacama Cosmology Telescope (ACT) during its 2008 season. ACT measures fluctuations at scales 500<l<10000. We fit a model for the lensed CMB, Sunyaev-Zel'dovich (SZ), and foreground contribution to the 148 GHz and 218 GHz power spectra, including thermal and kinetic SZ, Poisson power from radio and infrared point sources, and clustered power from infrared point sources. The power from thermal and kinetic SZ at 148 GHz is estimated to be B_3000 = 6.8+-2.9 uK^2, where B_l=l(l+1)C_l/2pi. We estimate primary cosmological parameters from the 148 GHz spectrum, marginalizing over SZ and source power. The LCDM cosmological model is a good fit to the data, and LCDM parameters estimated from ACT+WMAP are consistent with the 7-year WMAP limits, with scale invariant n_s = 1 excluded at 99.7% CL (3sigma). A model with no CMB lensing is disfavored at 2.8sigma. By measuring the third to seventh acoustic peaks, and probing the Silk damping regime, the ACT data improve limits on cosmological parameters that affect the small-scale CMB power. The ACT data combined with WMAP give a 6sigma detection of primordial helium, with Y_P = 0.313+-0.044, and a 4sigma detection of relativistic species, assumed to be neutrinos, with Neff = 5.3+-1.3 (4.6+-0.8 with BAO+H0 data). From the CMB alone the running of the spectral index is constrained to be dn/dlnk = -0.034 +- 0.018, the limit on the tensor-to-scalar ratio is r<0.25 (95% CL), and the possible contribution of Nambu cosmic strings to the power spectrum is constrained to string tension Gmu<1.6 \times 10^-7 (95% CL).Comment: 20 pages, 13 figures. Submitted to ApJ. This paper is a companion to Hajian et al. (2010) and Das et al. (2010

A Cryogenic Silicon Interferometer for Gravitational-wave Detection

The detection of gravitational waves from compact binary mergers by LIGO has opened the era of gravitational wave astronomy, revealing a previously hidden side of the cosmos. To maximize the reach of the existing LIGO observatory facilities, we have designed a new instrument that will have 5 times the range of Advanced LIGO, or greater than 100 times the event rate. Observations with this new instrument will make possible dramatic steps toward understanding the physics of the nearby universe, as well as observing the universe out to cosmological distances by the detection of binary black hole coalescences. This article presents the instrument design and a quantitative analysis of the anticipated noise floor

Adding tree rings to North America's National Forest Inventories: an essential tool to guide drawdown of atmospheric CO2

Tree-ring time series provide long-term, annually resolved information on the growth of trees. When sampled in a systematic context, tree-ring data can be scaled to estimate the forest carbon capture and storage of landscapes, biomes, and-ultimately-the globe. A systematic effort to sample tree rings in national forest inventories would yield unprecedented temporal and spatial resolution of forest carbon dynamics and help resolve key scientific uncertainties, which we highlight in terms of evidence for forest greening (enhanced growth) versus browning (reduced growth, increased mortality). We describe jump-starting a tree-ring collection across the continent of North America, given the commitments of Canada, the United States, and Mexico to visit forest inventory plots, along with existing legacy collections. Failing to do so would be a missed opportunity to help chart an evidence-based path toward meeting national commitments to reduce net greenhouse gas emissions, urgently needed for climate stabilization and repair.Published versio

Mitogen-induced stimulation and suppression of erythroid burst promoting activity production by human mononuclear cells

Exposure of human peripheral blood mononuclear cells or highly enriched monocytes to various plant lectins substantially alters their production of erythroid burst promoting activity (BPA). Neither unstimulated, nor mitogen stimulated, enriched T lymphocytes produced demonstrable BPA. Each of the lectins tested resulted in a different pattern of alteration of BPA production by mononuclear cells. Increasing concentrations of phytohaemagglutinin (PHA) caused a progressive increase in BPA production up to a plateau level at concentrations above 0·25–0·5 Μ1/ml. Concanavalin A (Con A) at concentrations of 0·05–0·1 Μg/ml stimulated BPA production, but Con A concentrations > 1 Μg/ml never augmented BPA production by mononuclear cells. Pokeweed mitogen inhibited BPA production by mononuclear cells in a concentration-dependent manner. Since PHA and Con A can bind to and stimulate both monocytes/macrophages and T lymphocytes, some production of BPA by stimulated T cells in the presence of monocytes cannot be ruled out. Earlier studies demonstrated that T cells augment monocyte production of BPA. Thus, monocyte–T cell interactions, as well as activation of monocytes and perhaps lymphocytes, play an important role in regulation of BPA production in vitro .Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/73808/1/j.1365-2141.1983.tb01232.x.pd