Kinetic Mechanism of Human Histidine Triad Nucleotide Binding Protein 1

Human histidine triad nucleotide binding protein 1 (hHint1) is a member of a ubiquitous and ancient branch of the histidine triad protein superfamily. hHint1 is a homodimeric protein that catalyzes the hydrolysis of model substrates, phosphoramidate and acyl adenylate, with a high efficiency. Recently, catalytically inactive hHint1 has been identified as the cause of inherited peripheral neuropathy [Zimon, M., et al. (2012) Nat. Genet. 44, 1080-1083]. We have conducted the first detailed kinetic mechanistic studies of hHint1 and have found that the reaction mechanism is consistent with a double-displacement mechanism, in which the active site nucleophile His112 is first adenylylated by the substrate, followed by hydrolysis of the AMP-enzyme intermediate. A transient burst phase followed by a linear phase from the stopped-flow fluorescence assay indicated that enzyme adenylylation was faster than the subsequent intermediate hydrolysis and product release. Solvent viscosity experiments suggested that both chemical transformation and diffusion-sensitive events (product release or protein conformational change) limit the overall turnover. The catalytic trapping experiments and data simulation indicated that the true koff rate of the final product AMP is unlikely to control the overall kcat. Therefore, a protein conformational change associated with product release is likely rate-limiting. In addition, the rate of Hint1 adenylylation was found to be dependent on two residues with pKa values of 6.5 and 8, with the former pKa agreeing well with the nuclear magnetic resonance titration results for the pKa of the active site nucleophile His112. In comparison to the uncatalyzed rates, hHint1 was shown to enhance acyl-AMP and AMP phosphoramidate hydrolysis by 10(6)-10(8)-fold. Taken together, our analysis indicates that hHint1 catalyzes the hydrolysis of phosphoramidate and acyl adenylate with high efficiency, through a mechanism that relies on rapid adenylylation of the active residue, His112, while being partially rate-limited by intermediate hydrolysis and product release associated with a conformational change. Given the high degree of sequence homology of Hint proteins across all kingdoms of life, it is likely that their kinetic and catalytic mechanisms will be similar to those elucidated for hHint1

Public health campaigns and obesity - a critique

<p>Abstract</p> <p>Background</p> <p>Controlling obesity has become one of the highest priorities for public health practitioners in developed countries. In the absence of safe, effective and widely accessible high-risk approaches (e.g. drugs and surgery) attention has focussed on community-based approaches and social marketing campaigns as the most appropriate form of intervention. However there is limited evidence in support of substantial effectiveness of such interventions.</p> <p>Discussion</p> <p>To date there is little evidence that community-based interventions and social marketing campaigns specifically targeting obesity provide substantial or lasting benefit. Concerns have been raised about potential negative effects created by a focus of these interventions on body shape and size, and of the associated media targeting of obesity.</p> <p>Summary</p> <p>A more appropriate strategy would be to enact high-level policy and legislative changes to alter the obesogenic environments in which we live by providing incentives for healthy eating and increased levels of physical activity. Research is also needed to improve treatments available for individuals already obese.</p

Reducing the environmental impact of surgery on a global scale: systematic review and co-prioritization with healthcare workers in 132 countries

Abstract Background Healthcare cannot achieve net-zero carbon without addressing operating theatres. The aim of this study was to prioritize feasible interventions to reduce the environmental impact of operating theatres. Methods This study adopted a four-phase Delphi consensus co-prioritization methodology. In phase 1, a systematic review of published interventions and global consultation of perioperative healthcare professionals were used to longlist interventions. In phase 2, iterative thematic analysis consolidated comparable interventions into a shortlist. In phase 3, the shortlist was co-prioritized based on patient and clinician views on acceptability, feasibility, and safety. In phase 4, ranked lists of interventions were presented by their relevance to high-income countries and low–middle-income countries. Results In phase 1, 43 interventions were identified, which had low uptake in practice according to 3042 professionals globally. In phase 2, a shortlist of 15 intervention domains was generated. In phase 3, interventions were deemed acceptable for more than 90 per cent of patients except for reducing general anaesthesia (84 per cent) and re-sterilization of ‘single-use’ consumables (86 per cent). In phase 4, the top three shortlisted interventions for high-income countries were: introducing recycling; reducing use of anaesthetic gases; and appropriate clinical waste processing. In phase 4, the top three shortlisted interventions for low–middle-income countries were: introducing reusable surgical devices; reducing use of consumables; and reducing the use of general anaesthesia. Conclusion This is a step toward environmentally sustainable operating environments with actionable interventions applicable to both high– and low–middle–income countries

Comparison of Optical Transition Radiation Simulations and Theory

The majority of optical diagnostics currently used will not stand up to the requirements of the next generation of particle accelerators. Current methodologies need innovation to be able to reach the sub-micrometre resolution and sensitivity that will be required. One technique that has the potential to meet these requirements is optical transition radiation (OTR) imaging. A new algorithm is proposed which incorporates OTR theory, optical effects and beam distribution. This algorithm takes an existing method used for beam imaging and pushes the limits resolution beyond that normally attainable. In doing so, it can provide a reliable and economical diagnostic for future accelerators. A discussion on further applications of the algorithm is also presented

Optical Effects in High Resolution and High Dynamic Range Beam Imaging Systems

Optical systems are used to transfer light in beam diagnostics for a variety of imaging applications. The effect of the point spread function (PSF) of these optical systems on the resulting measurements is often approximated or misunderstood. It is imperative that the optical PSF is independently characterised, as this can severely impede the attainable resolution of a diagnostic measurement. A high quality laser and specially chosen optics have been used to generate an intense optical point source in order to accomplish such a characterisation. The point source was used to measure the PSFs of various electron-beam imaging systems. These systems incorporate a digital micro-mirror array, which was used to produce very high (>105) dynamic range images. The PSF was measured at each intermediary image plane of the optical system; enabling the origin of any perturbations to the PSF to be isolated and potentially mitigated. One of the characterised systems has been used for optical transition radiation (OTR) measurements of an electron beam at KEK-ATF2 (Tsukuba, Japan). This provided an application of this process to actively improve the resolution of the beam imaging system. Presented here are the results of our measurements and complementary simulations carried out using Zemax Optical Studio

High Resolution and Dynamic Range Characterisation of Beam Imaging Systems

Any imaging system requires the use of various optical components to transfer the light from the source, e.g. optical radiation generated by a charged particle beam, to the sensor. The impact of the transfer optics on the image resolution is often not well known. To improve this situation, the point spread function (PSF) of the optical system must be measured, preferably, with high dynamic range. For this purpose we have created an intense, small (~ 1 μm) point source using a high quality laser and special focusing optics; and introduced a digital micro-mirror array in the optical system to substantially increase its dynamic range. The PSFs of optical systems that are currently being developed for high resolution, high dynamic range beam imaging using optical transition and diffraction radiation are measured and compared to Zemax simulations. The goal of these studies is to systematically understand and mitigate any ill effects on the PSF due to aberrations, diffraction and misalignment of the components of the imaging system. We present the results of our measurements and simulations

Cherenkov Radiation in Optical Fibres as a Versatile Machine Protection System in Particle Accelerators.

Machine protection systems in high power particle accelerators are crucial. They can detect, prevent, and respond to events which would otherwise cause damage and significant downtime to accelerator infrastructure. Current systems are often resource heavy and operationally expensive, reacting after an event has begun to cause damage; this leads to facilities only covering certain operational modes and setting lower limits on machine performance. Presented here is a new type of machine protection system based upon optical fibres, which would be complementary to existing systems, elevating existing performance. These fibres are laid along an accelerator beam line in lengths of ∼100 m, providing continuous coverage over this distance. When relativistic particles pass through these fibres, they generate Cherenkov radiation in the optical spectrum. This radiation propagates in both directions along the fibre and can be detected at both ends. A calibration based technique allows the location of the Cherenkov radiation source to be pinpointed to within 0.5 m with a resolution of 1 m. This measurement mechanism, from a single device, has multiple applications within an accelerator facility. These include beam loss location monitoring, RF breakdown prediction, and quench prevention. Detailed here are the application processes and results from measurements, which provide proof of concept for this device for both beam loss monitoring and RF breakdown detection. Furthermore, highlighted are the current challenges for future innovation

Linearity and Response Time of the LHC Diamond Beam Loss Monitors in the CLEAR Beam Test Facility at CERN

Chemical Vapour Deposition (CVD) diamond detectors have been tested during the Run 2 operation period (2015-2018) as fast beam loss monitors for the Beam Loss Monitoring (BLM) system of the Large Hadron Collider (LHC) at CERN. However, the lack of raw data recorded during this operation period restrains our ability to perform a deep analysis of their signals. For this reason, a test campaign was carried out at the CLEAR beam test facility at CERN with the aim of studying the linearity and response time of the diamond detectors against losses from electron beams of different intensities. The signal build-up from multi-bunched electron beams was also analyzed. The conditions and procedures of the test campaign are explained, as well as the most significant results obtained

Recent AWAKE Diagnostics Development and Operational Results

The Advanced Wakefield Experiment (AWAKE) at CERN investigates the Plasma-Wakefield acceleration of electrons driven by a relativistic proton bunch. After successfully demonstrating the acceleration process in the AWAKE Run 1, the experiment has now started the Run 2. The AWAKE Run 2 consists of several experimental periods that aim to demonstrate the feasibility of the AWAKE concept beyond the acceleration experiment, showing its feasibility as accelerator for particle physics application. As part of these developments, a dramatic effort in improving the AWAKE instrumentation is sustained. This contribution reports on the current developments of the instrumentation pool upgrade, including the digital camera system for transverse beam profile measurement, beam halo measurement and the spectrometer upgrade studies. The studies on the development of high-frequency beam position monitors are also described