A bidirectional fluorescent two-hybrid system for monitoring protein–protein interactions

Two-hybrid systems have been the cornerstone of research into protein–protein interactions, but these systems typically rely on life/death reporters that put additional selective pressure on the host organism, and potentially lead to false positives. Here we report a bidirectional fluorescence-based bacterial two- hybrid system that enables both the association and dissociation of a given protein–protein interaction to be monitored. The functionality of this system and its compatibility with FACS screening are demon- strated in the forward and reverse direction using known interacting protein-partners and their cyclic peptide inhibitors. The reported fluorescent two-hybrid system may be used in the forward direction for the identification of interacting protein partners, or as a reverse two-hybrid system for the high- throughput identification of protein–protein interaction inhibitors

Fully Self-consistent RPA description of the many level pairing model

The Self-Consistent RPA (SCRPA) equations in the particle-particle channel are solved without any approximation for the picket fence model. The results are in excellent agreement with the exact solutions found with the Richardson method. Particularly interesting features are that screening corrections reverse the sign of the interaction and that SCRPA yields the exact energies in the case of two levels with two particles.Comment: 37 pages, 1 figure and 17 table

Bank capital regulation with and without state-contingent penalties

A moral hazard model with exogenous bank franchise value is used to analyze bank capital regulation. Banks choose their capital structure as well as the riskiness and mean of their portfolio. The portfolio mean is determined by the level of costly screening. Screening and portfolio risk are private information, so there are two dimensions to the moral hazard problem. Deposit insurance gives banks an incentive to hold less capital, and to choose a higher-risk, lower-mean portfolio. To mitigate these incentives, capital requirements with and without ex post fines are studied. We find an endogenous reverse mean-variance trade-off in banks' portfolios. Prudent banks choose high-screening, low-risk portfolios and are virtually self regulating. Imprudent banks choose low-screening, high-risk portfolios. Without state-contingent penalties, optimal capital regulations are often V-shaped in bank franchise value. Adding state-contingent regulation can significantly lower capital requirements. Optimal state-contingent regulations are characterized by fines on extreme right-hand-tail returns.Bank capital

Comparison of methods for in-house screening of HLA*B57:01 to prevent abacavir hypersensitivity in HIV-1 care

Abacavir is a nucleoside reverse transcriptase inhibitor used as part of combination antiretroviral therapy in HIV-1-infected patients. Because this drug can cause a hypersensitivity reaction that is correlated with the presence of the HLA-B*57:01 allotype, screening for the presence of HLA-B*57:01 is recommended before abacavir initiation. Different genetic assays have been developed for HLA-B*57:01 screening, each with specific sensitivity, turnaround time and assay costs. Here, a new real-time PCR (qPCR) based analysis is described and compared to sequence specific primer PCR with capillary electrophoresis (SSP PCR CE) on 149 patient-derived samples, using sequence specific oligonucleotide hybridization combined with high resolution SSP PCR as gold standard. In addition to these PCR based methods, a complementary approach was developed using flow cytometry with an HLA-B17 specific monoclonal antibody as a pre-screening assay to diminish the number of samples for genetic testing. All three assays had a maximum sensitivity of >99. However, differences in specificity were recorded, i.e. 84.3%, 97.2% and >99% for flow cytometry, qPCR and SSP PCR CE respectively. Our data indicate that the most specific and sensitive of the compared methods is the SSP PCR CE. Flow cytometry pre-screening can substantially decrease the number of genetic tests for HLA-B*57:01 typing in a clinical setting

Fuzzy virtual ligands for virtual screening

A new method to bridge the gap between ligand and receptor-based methods in virtual screening (VS) is presented. We introduce a structure-derived virtual ligand (VL) model as an extension to a previously published pseudo-ligand technique [1]: LIQUID [2] fuzzy pharmacophore virtual screening is combined with grid-based protein binding site predictions of PocketPicker [3]. This approach might help reduce bias introduced by manual selection of binding site residues and introduces pocket shape information to the VL. It allows for a combination of several protein structure models into a single "fuzzy" VL representation, which can be used to scan screening compound collections for ligand structures with a similar potential pharmacophore. PocketPicker employs an elaborate grid-based scanning procedure to determine buried cavities and depressions on the protein's surface. Potential binding sites are represented by clusters of grid probes characterizing the shape and accessibility of a cavity. A rule-based system is then applied to project reverse pharmacophore types onto the grid probes of a selected pocket. The pocket pharmacophore types are assigned depending on the properties and geometry of the protein residues surrounding the pocket with regard to their relative position towards the grid probes. LIQUID is used to cluster representative pocket probes by their pharmacophore types describing a fuzzy VL model. The VL is encoded in a correlation vector, which can then be compared to a database of pre-calculated ligand models. A retrospective screening using the fuzzy VL and several protein structures was evaluated by ten fold cross-validation with ROC-AUC and BEDROC metrics, obtaining a significant enrichment of actives. Future work will be devoted to prospective screening using a novel protein target of Helicobacter pylori and compounds from commercial providers

Strategies for increasing uptake of cervical screening

Practice nurses can help get cervical screening back on the rise by understanding the reasons for non-attendance and then tailoring the primary care provision to encourage more women to have the test, writes Aine Kothari Cervical screening uptake is declining throughout the UK. A knowledge of the reasons for non-attendance can be used to tailor primary care provision of cervical screening to encourage women to respond to their screening invitation. Cervical screening saves lives by early detection and treatment of pre-cancerous abnormalities. The cervical screening procedure takes approximately 15 minutes and the result is generally available within two weeks of the test. This article discusses the barriers to attendance and the strategies that might be adopted to reverse the downward trend in screening uptake

A protocol for a systematic review of clinical guidelines and published systematic reviews on the early detection of oral cancer

Background: The predicted increase in incidence of oral cavity cancer (OCC) coupled with high mortality and poor prognosis – particularly when diagnosed at a late/advanced stage – highlights the need for prevention and early detection/screening to reverse these trends. Dental healthcare professionals in primary care settings have a pivotal role in this effort. Aim: The aim of this protocol is to detail the process for assessing the evidence for the best practice and methods of early detection/screening for OCC in primary care dental settings by undertaking a systematic review of global clinical guidelines and published systematic reviews. Method: Searches for clinical guidelines and systematic reviews will be conducted in the following databases: Cochrane library, Medical Literature Analysis and Retrieval System Online (Ovid), Excerpta Medical dataBASE, PubMed, Turning Research into Practice, SCOPUS and Web of Science Core Collection. Our search will extend to include Google Scholar and international professional organizations/associations websites. In addition, we will handsearch the bibliographies and undertake citation searches of the selected papers. Quality appraisal will be undertaken using the Appraisal of Guidelines for Research and Evaluation version II instrument for the clinical guidelines and both A MeaSurement Tool to Assess Systematic Reviews and Risk of Bias in Systematic Reviews tools for the systematic reviews. A narrative synthesis approach will be used to assess the evidence of extracted data, primarily taking account of quality appraisal and recency of publication. Discussion: The synthesis of evidence will determine best practice for OCC early detection/screening by primary care dental healthcare professionals and will evaluate the relationship between clinical guidelines and the evidence base available from systematic reviews in this area

Improved Screening of cDNAs Generated by mRNA Differential Display Enables the Selection of True Positives and the Isolation of Weakly Expressed Messages

The high percentage of false positives generated by differential display (as high as 85%) has previously limited the potential of the method. This report describes an efficient methodology that enables false positives to be discarded prior to cloning, via reverse Northern analysis. This first step of the screening also allows the detection of putative lowabundance differential clones. Following cloning, a second reverseNorthern combined with partial DNA sequencing and RT-PCR detection allows isolation of all differential cDNAs including very lowabundance clones. Use of the sequential screening procedure described here led to the isolation of novel tomato genes responding to the plant hormone ethylene while minimising labor and materials input

The genome of the protozoan parasite Cystoisospora suis and a reverse vaccinology approach to identify vaccine candidates

Vaccine development targeting protozoan parasites remains challenging, partly due to the complex interactions between these eukaryotes and the host immune system. Reverse vaccinology is a promising approach for direct screening of genome sequence assemblies for new vaccine candidate proteins. Here, we applied this paradigm to Cystoisospora suis, an apicomplexan parasite that causes enteritis and diarrhea in suckling piglets and economic losses in pig production worldwide. Using Next Generation Sequencing we produced an ∼84 Mb sequence assembly for the C. suis genome, making it the first available reference for the genus Cystoisospora. Then, we derived a manually curated annotation of more than 11,000 protein-coding genes and applied the tool Vacceed to identify 1,168 vaccine candidates by screening the predicted C. suis proteome. To refine the set of candidates, we looked at proteins that are highly expressed in merozoites and specific to apicomplexans. The stringent set of candidates included 220 proteins, among which were 152 proteins with unknown function, 17 surface antigens of the SAG and SRS gene families, 12 proteins of the apicomplexan-specific secretory organelles including AMA1, MIC6, MIC13, ROP6, ROP12, ROP27, ROP32 and three proteins related to cell adhesion. Finally, we demonstrated in vitro the immunogenic potential of a C. suis-specific 42 kDa transmembrane protein, which might constitute an attractive candidate for further testing