Structural interrogation of phosphoproteome identified by mass spectrometry reveals allowed and disallowed regions of phosphoconformation

High-throughput mass spectrometric (HT-MS) study is the method of choice for monitoring global changes in proteome. Data derived from these studies are meant for further validation and experimentation to discover novel biological insights. Here we evaluate use of relative solvent accessible surface area (rSASA) and DEPTH as indices to assess experimentally determined phosphorylation events deposited in PhosphoSitePlus. Based on accessibility, we map these identifications on allowed (accessible) or disallowed (inaccessible) regions of phosphoconformation. Surprisingly a striking number of HT- MS/MS derived events (1461/5947 sites or 24.6%) are present in the disallowed region of conformation. By considering protein dynamics, autophosphorylation events and/or the sequence specificity of kinases, 13.8% of these phosphosites can be moved to the allowed region of conformation. We also demonstrate that rSASA values can be used to increase the confidence of identification of phosphorylation sites within an ambiguous MS dataset. While MS is a stand-alone technique for the identification of vast majority of phosphorylation events, identifications within disallowed region of conformation will benefit from techniques that independently probe for phosphorylation and protein dynamics. Our studies also imply that trapping alternate protein conformations may be a viable alternative to the design of inhibitors against mutation prone drug resistance kinases

SUSPENSION POLICY FOR CLOUD-BASED VIRTUAL MACHINES

Techniques are described herein for obtaining usage metrics of Virtual Machines (VMs) over a number of days. This data may be used to automatically identify periodic (e.g., daily) blocks of time when groups of VMs can be suspended. Unlike typical scenarios in which VMs in a cloud-based environment stay up irrespective of whether the VM is being used, these techniques avoid unnecessary cost to the client

Chemisorption and FTIR study of bimetallic Pt---Au/SiO₂catalysts

Pt/SiO₂, Au/SiO₂, and bimetallic Pt---Au/SiO₂ catalysts were prepared by incipient wetness impregnation of nonporous SiO₂. The catalysts were characterized after reduction in H₂ by static volumetric chemisorption and infrared spectroscopy. For the monometallic and the bimetallic catalysts, H₂, O₂, and CO were used as adsorbates at room temperature. Additionally, for the monometallic Au/SiO₂ catalyst, OZ adsorption at 473 K was also used. Infrared spectra of adsorbed CO were obtained on all catalysts. Addition of gold decreased the uptake of all three adsorbates at room temperature, without significantly influencing the relative amounts of weakly held adsorbed species which could be removed upon pumping for 30 min. Agreeing with CO chemisorption data, the total integrated intensity of the linear CO band decreased with increasing Au content. The IR results indicated a predominantly geometric effect of Au causing a shift of the linear CO band to lower wavenumbers. The shift could be rationalized in terms of decreased dipole-dipole coupling of adsorbed CO species. The results are discussed within the context of a previous investigation using electron microscopy, TPR, and reactivity data. Portions of the bimetallic catalysts were subjected to three high-temperature oxidation/reduction cycles and characterized by static chemisorption of H₂ to investigate the effect of this thermal treatment on Pt dispersion.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/28743/1/0000573.pd

Evidence-based practice at Patan Academy of Health Sciences, Nepal: knowledge, attitude, behavior and barriers

Introduction: Evidence based practice (EBP) is the use of current best evidence in making decisions about the care of individual patients. The aim of this study was to explore the practices, attitudes, knowledge and perceived barriers in relation to EBP among faculty members at Patan Academy of Health Sciences (PAHS), Nepal. Methods: This was a cross-sectional survey study. Participants were faculty members (both clinical and non-clinical) at PAHS. The main outcomes were attitude towards the usefulness of EBP, the frequency of use of EBP in practice, and the barriers to carrying out EBP. Results: One hundred and seventy-two faculty members participated in this survey. Ninety five percent knew it was used for clinical decision making and the majority used it for teaching. Senior faculty spent more time on searching for evidence and reading compared to junior faculty. There was no difference between clinical and non-clinical departments. The main barriers to EBP were inadequate teaching learning support by 49% (85/172) followed by inadequate time 30% (52/172). Conclusions: The study showed that faculty at PAHS had a generally positive attitude towards EBP. EBP could be better facilitated by proper teaching and allocation of dedicated time. Keywords: evidence-based practice, faculties, health sciences, attitude- knowledge-behavio

Development of a Cytosensor for the Detection of Fusarium Oxysporum - A Functional Approach Towards Bioanalytical Applications

The development of bio-analytical methods for monitoring microorganisms have created opportunities for applications in biosensors, bioprocess monitoring, assessment of cell signalling, analysis of drug responses, among several others. The voltammetric sensing system employed for studying the electrode behavior of the fungus Fusarium oxysporum comprised working (gold) electrode (0.2 cm2 ) platinum as counter electrode (0.2 cm2 ) and a saturated calomel as the reference, where the electrochemical response corresponded to the growth phases (lag, log, stationary and decline) of the fungus. The electrochemical method based on voltammetric response matched well with the response obtained through conventional methodology, where the dry weight of the fungus is estimated against time. The peak potential is a function of scan rate, which is one of the characteristic features of a totally irreversible electrode process. It is important to mention here that this dependence is true regardless of reversibility for any diffusing redoxactive species. The proposed electrochemical method is less cumbersome and more accurate. Furthermore, the proposed electrochemical method captures the decline phase of fungal growth, which is generally difficult using the conventional method of assessment of the growth curve. Further experiments confirm that the anodic peaks were not due to the biomass or the fungal spores and only due to the extracellular metabolites. However, at this stage it is difficult to exactly determine the metabolite or the group of metabolites that are responsible for the anodic peak. In conclusion this cytosensor is capable of accurately and rapidly quantifying fungi with Fusarium oxysporum as a model organism

Upgrading Lignocellulosic Products to Drop-In Biofuels via Dehydrogenative Cross-Coupling and Hydrodeoxygenation Sequence

Life-cycle analysis (LCA) allows the scientific community to identify the sources of greenhouse gas (GHG) emissions of novel routes to produce renewable fuels. Herein, we integrate LCA into our investigations of a new route to produce drop-in diesel/jet fuel by combining furfural, obtained from the catalytic dehydration of lignocellulosic pentose sugars, with alcohols that can be derived from a variety of bio- or petroleum-based feedstocks. As a key innovation, we developed recyclable transition-metal-free hydrotalcite catalysts to promote the dehydrogenative cross-coupling reaction of furfural and alcohols to give high molecular weight adducts via a transfer hydrogenation- aldol condensation pathway. Subsequent hydrodeoxygenation of adducts over Pt/NbOPO4 yields alkanes. Implemented in a Brazilian sugarcane biorefinery such a process could result in a 53-79% reduction in life-cycle GHG emissions relative to conventional petroleum fuels and provide a sustainable source of low carbon diesel/jet fuel

Non-coding nucleotides and amino acids near the active site regulate peptide deformylase expression and inhibitor susceptibility in Chlamydia trachomatis

Chlamydia trachomatis, an obligate intracellular bacterium, is a highly prevalent human pathogen. Hydroxamic-acid-based matrix metalloprotease inhibitors can effectively inhibit the pathogen both in vitro and in vivo, and have exhibited therapeutic potential. Here, we provide genome sequencing data indicating that peptide deformylase (PDF) is the sole target of the inhibitors in this organism. We further report molecular mechanisms that control chlamydial PDF (cPDF) expression and inhibition efficiency. In particular, we identify the σ66-dependent promoter that controls cPDF gene expression and demonstrate that point mutations in this promoter lead to resistance by increasing cPDF transcription. Furthermore, we show that substitution of two amino acids near the active site of the enzyme alters enzyme kinetics and protein stability