In Vitro Evolution of Allergy Vaccine Candidates, with Maintained Structure, but Reduced B Cell and T Cell Activation Capacity

Allergy and asthma to cat (Felis domesticus) affects about 10% of the population in affluent countries. Immediate allergic symptoms are primarily mediated via IgE antibodies binding to B cell epitopes, whereas late phase inflammatory reactions are mediated via activated T cell recognition of allergen-specific T cell epitopes. Allergen-specific immunotherapy relieves symptoms and is the only treatment inducing a long-lasting protection by induction of protective immune responses. The aim of this study was to produce an allergy vaccine designed with the combined features of attenuated T cell activation, reduced anaphylactic properties, retained molecular integrity and induction of efficient IgE blocking IgG antibodies for safer and efficacious treatment of patients with allergy and asthma to cat. The template gene coding for rFel d 1 was used to introduce random mutations, which was subsequently expressed in large phage libraries. Despite accumulated mutations by up to 7 rounds of iterative error-prone PCR and biopanning, surface topology and structure was essentially maintained using IgE-antibodies from cat allergic patients for phage enrichment. Four candidates were isolated, displaying similar or lower IgE binding, reduced anaphylactic activity as measured by their capacity to induce basophil degranulation and, importantly, a significantly lower T cell reactivity in lymphoproliferative assays compared to the original rFel d 1. In addition, all mutants showed ability to induce blocking antibodies in immunized mice.The approach presented here provides a straightforward procedure to generate a novel type of allergy vaccines for safer and efficacious treatment of allergic patients

Family Business Restructuring:A Review and Research Agenda

Although business restructuring occurs frequently and it is important for the prosperity of family firms across generations, research on family firms has largely evolved separately from research on business restructuring. This is a missed opportunity, since the two domains are complementary, and understanding the context, process, content, and outcome dimensions is relevant to both research streams. We address this by examining the intersection between research on business restructuring and family firms to improve our knowledge of each area and inform future research. To achieve this goal, we review and organize research across different dimensions to create an integrative framework. Building on current research, we focus on 88 studies at the intersection of family firm and business restructuring research to develop a model that identifies research needs and suggests directions for future research

Air Pollution Dynamics and the Need for Temporally Differentiated Road Pricing

In this paper we investigate the effects of the temporal variation of pollution dispersion, traffic flows and vehicular emissions on pollution concentration and illustrate the need for temporally differentiated road pricing through an application to the case of the congestion charge in Stockholm, Sweden. By accounting explicitly for the role of pollution dispersion on optimal road pricing, we allow for a more comprehensive view of the economy-ecology interactions at stake, showing that price differentiation is an optimal response to the physical environment. Most congestion charges in place incorporate price bands to mitigate congestion. Our analysis indicates that, to ensure compliance with air quality standards, such price variations should also be a response to limited pollution dispersion. © 2015 Elsevier Ltd

Urban NO2 and NO Pollution in Relation to the North Atlantic Oscillation NAO

The North Atlantic Oscillation (NAO), a measure of the strength of the zonal wind across the North Atlantic Ocean, strongly influences weather conditions in NW Europe, e.g. temperature, precipitation and wind, especially during winter. It was hypothesised that elevated concentrations of nitrogen oxides in Gothenburg would be enhanced during negative NAO index (NAOI) conditions, representing more anticyclonic weather situations and thus leading to limited air mixing in the urban atmosphere, than situations with NAOI > 0. Hourly wintertime (December–February) concentrations (1997–2006) of NO2, NO, air pressure, temperature and wind direction from an urban rooftop (30 m above street level) in the centre of the City of Gothenburg were analysed in relation to NAOI. Air pressure, the average concentration of nitrogen oxides (NOx = NO2 + NO), as well as the fraction of hourly NO2 and NO concentrations exceeding 90 μg m−3 and the fraction of daily NO concentrations exceeding 60 μg m−3, were significantly and negatively related to NAOI. Air temperature was positively correlated with NAOI. Southerly and westerly winds were more common in months with positive NAOI, while easterly and northerly winds were overrepresented in months with negative NAOI. High pollution concentrations dominantly occurred in situations with northerly and easterly wind directions. High NO2 and NO concentrations were associated with negative NAOI, especially in the morning when the traffic rush coincided with restricted air mixing. Over the ten-year period there were trends for more negative NAOI and increased time fractions with hourly NO2 concentrations exceeding 90 μg m−3. The conclusion of this study is that a climate shift towards higher or lower NAOI has the potential to significantly influence urban air pollution in North-West Europe, and thus the possibility to reach air quality standards, even if emissions remain constant

Directed evolution of a type I antifreeze protein expressed in Escherichia coli with sodium chloride as selective pressure and its effect on antifreeze tolerance

Both freezing tolerance and NaCl tolerance are improved when antifreeze proteins are expressed as fusion proteins with two domains of staphylococcal protein A (SPA) in Escherichia coli. To characterize these properties further we created a randomly mutated expression library in E. coli, based on the winter flounder antifreeze protein HPLC-8 component gene. Low-fidelity PCR products of this gene were fused to the spa gene encoding two domains of the SPA. The library was screened for enhanced NaCl tolerance and four clones were selected. The freezing tolerance of each of the selected clones was enhanced to varying extents. DNA sequencing of the isolated mutants revealed that the amphiphilic properties of the native antifreeze protein were essentially conserved. Furthermore, by studying the primary sequence of the randomly mutated clones, in comparison with the degree of freezing tolerance, we have identified clues which help in understanding the relationship between salt and freezing tolerance

Location and nucleotide sequence of frdB, the gene coding for the iron-sulphur protein subunit of the fumarate reductase of Escherichia coli

The frdB gene, encoding the iron-sulphur protein subunit of fumarate reductase, has been located and its complete nucleotide sequence determined. The identity of the gene was confirmed by protein chemical studies and determination of the NH2-terminal sequence of the FrdB protein. The frdB gene is situated distal to and partially overlapped by frdA which codes for the flavoprotein subunit of the reductase. Its reading frame contains 244 codons and predicts a protein of Mr 27092. In composition, the FrdB protein is strikingly similar to the corresponding subunit of the related flavoenzyme, succinate dehydrogenase. Analysis of the protein's primary structure revealed several features characteristic of iron-sulphur proteins