Substrate specificity of bacterial oligosaccharyltransferase suggests a common transfer mechanism for the bacterial and eukaryotic systems

The PgIB oligosaccharyltransferase (OTase) of Campylobacter jejuni can be functionally expressed in Escherichia coli, and its relaxed oligosaccharide substrate specificity allows the transfer of different glycans from the lipid carrier undecaprenyl pyrophosphate to an acceptor protein. To investigate the substrate specificity of PgIB, we tested the transfer of a set of lipid-linked polysaccharides in E. coli and Salmonella enterica serovar Typhimurium. A hexose linked to the C-6 of the monosaccharide at the reducing end did not inhibit the transfer of the O antigen to the acceptor protein. However, PgIB required an acetamido group at the C-2. A model for the mechanism of PgIB involving this functional group was proposed. Previous experiments have shown that eukaryotic OTases have the same requirement, suggesting that eukaryotic and prokaryotic OTases catalyze the transfer of oligosaccharides by a conserved mechanism. Moreover, we demonstrated the functional transfer of the C. jejuni glycosylation system into S. enterica. The elucidation of the mechanism of action and the substrate specificity of PgIB represents the foundation for engineering glycoproteins that will have an impact on biotechnology

Paediatric obsessive-compulsive disorder and depressive symptoms: clinical correlates and CBT treatment outcomes.

Depression frequently co-occurs with paediatric obsessive-compulsive disorder (OCD), yet the clinical correlates and impact of depression on CBT outcomes remain unclear. The prevalence and clinical correlates of depression were examined in a paediatric specialist OCD-clinic sample (N = 295; Mean = 15 [7 - 18] years, 42 % female), using both dimensional (Beck Depression Inventory-youth; n = 261) and diagnostic (Development and Wellbeing Assessment; n = 127) measures of depression. The impact of depressive symptoms and suspected disorders on post-treatment OCD severity was examined in a sub-sample who received CBT, with or without SSRI medication (N = 100). Fifty-one per-cent of patients reported moderately or extremely elevated depressive symptoms and 26 % (95 % CI: 18 - 34) met criteria for a suspected depressive disorder. Depressive symptoms and depressive disorders were associated with worse OCD symptom severity and global functioning prior to CBT. Individuals with depression were more likely to be female, have had a psychiatric inpatient admission and less likely to be attending school (ps < 0.01). OCD and depressive symptom severity significantly decreased after CBT. Depressive symptoms and depressive disorders predicted worse post-treatment OCD severity (βs = 0.19 and 0.26, ps < 0.05) but became non-significant when controlling for pre-treatment OCD severity (βs = 0.05 and 0.13, ns). Depression is common in paediatric OCD and is associated with more severe OCD and poorer functioning. However, depression severity decreases over the course of CBT for OCD and is not independently associated with worse outcomes, supporting the recommendation for treatment as usual in the presence of depressive symptoms

Engineering Yarrowia lipolytica to Produce Glycoproteins Homogeneously Modified with the Universal Man3GlcNAc2 N-Glycan Core

Yarrowia lipolytica is a dimorphic yeast that efficiently secretes various heterologous proteins and is classified as “generally recognized as safe.” Therefore, it is an attractive protein production host. However, yeasts modify glycoproteins with non-human high mannose-type N-glycans. These structures reduce the protein half-life in vivo and can be immunogenic in man. Here, we describe how we genetically engineered N-glycan biosynthesis in Yarrowia lipolytica so that it produces Man3GlcNAc2 structures on its glycoproteins. We obtained unprecedented levels of homogeneity of this glycanstructure. This is the ideal starting point for building human-like sugars. Disruption of the ALG3 gene resulted in modification of proteins mainly with Man5GlcNAc2 and GlcMan5GlcNAc2 glycans, and to a lesser extent with Glc2Man5GlcNAc2 glycans. To avoid underoccupancy of glycosylation sites, we concomitantly overexpressed ALG6. We also explored several approaches to remove the terminal glucose residues, which hamper further humanization of N-glycosylation; overexpression of the heterodimeric Apergillus niger glucosidase II proved to be the most effective approach. Finally, we overexpressed an α-1,2-mannosidase to obtain Man3GlcNAc2 structures, which are substrates for the synthesis of complex-type glycans. The final Yarrowia lipolytica strain produces proteins glycosylated with the trimannosyl core N-glycan (Man3GlcNAc2), which is the common core of all complex-type N-glycans. All these glycans can be constructed on the obtained trimannosyl N-glycan using either in vivo or in vitro modification with the appropriate glycosyltransferases. The results demonstrate the high potential of Yarrowia lipolytica to be developed as an efficient expression system for the production of glycoproteins with humanized glycans

Cryo-electron microscopy of viruses

Thin vitrified layers of unfixed, unstained and unsupported virus suspensions can be prepared for observation by cryo-electron microscopy in easily controlled conditions. The viral particles appear free from the kind of damage caused by dehydration, freezing or adsorption to a support that is encountered in preparing biological samples for conventional electron microscopy. Cryo-electron microscopy of vitrified specimens offers possibilities for high resolution observations that compare favourably with any other electron microscopical method

Can burglary prevention be low-carbon and effective? Investigating the environmental performance of burglary prevention measures

There has been limited study to date on the environmental impacts of crime prevention measures. We address this shortfall by estimating the carbon footprint associated with the most widely used burglary prevention measures: door locks, window locks, burglar alarms, lighting and CCTV cameras. We compare these footprints with a measure of their effectiveness, the security protection factor, allowing us to identify those measures that are both low-carbon and effective in preventing burglary. Window locks are found to be the most effective and low-carbon measure available individually. Combinations of window locks, door locks, external and indoor lightings are also shown to be effective and low-carbon. Burglar alarms and CCTV do not perform as strongly, with low security against burglary and higher carbon footprints. This information can be used to help inform more sustainable choices of burglary prevention within households as well as for crime prevention product design

The Great American Crime Decline : Possible Explanations

This chapter examines the most important features of the crime decline in the United States during the 1990s-2010s but also takes a broader look at the violence declines of the last three centuries. The author argues that violent and property crime trends might have diverged in the 1990s, with property crimes increasingly happening in the online sphere and thus traditional property crime statistics not being reflective of the full picture. An important distinction is made between ‘contact crimes’ and crimes that do not require a victim and offender to be present in the same physical space. Contrary to the uncertainties engendered by property crime, the declines in violent (‘contact’) crime are rather general, and have been happening not only across all demographic and geographic categories within the United States but also throughout the developed world. An analysis of research literature on crime trends has identified twenty-four different explanations for the crime drop. Each one of them is briefly outlined and examined in terms of conceptual clarity and empirical support. Nine crime decline explanations are highlighted as the most promising ones. The majority of these promising explanations, being relative newcomers in the crime trends literature, have not been subjected to sufficient empirical scrutiny yet, and thus require further research. One potentially fruitful avenue for future studies is to examine the association of the most promising crime decline explanations with improvements in self-control

Virosome-Formulated Plasmodium falciparum AMA-1 & CSP Derived Peptides as Malaria Vaccine: Randomized Phase 1b Trial in Semi-Immune Adults & Children

BACKGROUND\ud \ud This trial was conducted to evaluate the safety and immunogenicity of two virosome formulated malaria peptidomimetics derived from Plasmodium falciparum AMA-1 and CSP in malaria semi-immune adults and children.\ud \ud METHODS\ud \ud The design was a prospective randomized, double-blind, controlled, age-deescalating study with two immunizations. 10 adults and 40 children (aged 5-9 years) living in a malaria endemic area were immunized with PEV3B or virosomal influenza vaccine Inflexal®V on day 0 and 90.\ud \ud RESULTS\ud \ud No serious or severe adverse events (AEs) related to the vaccines were observed. The only local solicited AE reported was pain at injection site, which affected more children in the Inflexal®V group compared to the PEV3B group (p = 0.014). In the PEV3B group, IgG ELISA endpoint titers specific for the AMA-1 and CSP peptide antigens were significantly higher for most time points compared to the Inflexal®V control group. Across all time points after first immunization the average ratio of endpoint titers to baseline values in PEV3B subjects ranged from 4 to 15 in adults and from 4 to 66 in children. As an exploratory outcome, we found that the incidence rate of clinical malaria episodes in children vaccinees was half the rate of the control children between study days 30 and 365 (0.0035 episodes per day at risk for PEV3B vs. 0.0069 for Inflexal®V; RR  = 0.50 [95%-CI: 0.29-0.88], p = 0.02).\ud \ud CONCLUSION\ud \ud These findings provide a strong basis for the further development of multivalent virosomal malaria peptide vaccines.\ud \ud TRIAL REGISTRATION\ud \ud ClinicalTrials.gov NCT00513669