Surface expression has attracted much recent interest, and it has been suggested for a variety of applications. Two such applications are whole-cell biocatalysis and the creation of live vaccines. For successful implementation of these applications there is a need for flexible surface expression systems that can yield a high level of expression with a variety of recombinant fusion proteins. The aim of this work was thus to create a surface expression system that would fulfil these requirements.   A novel surface expression system based on the AIDA-I autotransporter was created with the key qualities being are good, protein-independent detection of the expression through the presence of two epitope tags flanking the recombinant protein, and full modularity of the different components of the expression cassette. To evaluate the flexibility of this construct, 8 different model proteins with potential use as live-vaccines or biocatalysts were expressed and their surface expression levels were analysed.   Positive signals were detected for all of the studied proteins using antibody labelling followed by flow cytometric analysis, showing the functionality of the expression system. The ratio of the signal from the two epitope tags indicated that several of the studied proteins were present mainly in proteolytically degraded forms, which was confirmed by Western blot analysis of the outer membrane protein fraction. This proteolysis was suggested to be due to protein-dependent stalling of translocation intermediates in the periplasm, with indications that larger size and higher cysteine content had a negative impact on expression levels. Process design with reduced cultivation pH and temperature was used to increase total surface expression yield of one of the model proteins by 400 %, with a simultaneous reduction of proteolysis by a third. While not sufficient to completely remove proteolysis, this shows that process design can be used to greatly increase surface expression. Thus, it is recommended that future work combine this with engineering of the bacterial strain or the expression system in order to overcome the observed proteolysis and maximise the yield of surface expressed protein. / <p>QC 20130516</p>

No description available.

A relatively high percentage of porcine <i>Escherichia coli</i> isolates from cases associated with neonatal and post-weaning diarrhea are positive for the gene encoding the adhesin involved in diffuse adherence I (AIDA-I). This gene and its corresponding protein were first identified and characterized in <i>E. coli</i> strain 2787 isolated from human infantile diarrhea. Little is known about the role of the AIDA-I protein in pathogenesis of porcine enteric disease caused by AIDA-I positive E. coli and the properties of AIDA-I protein expressed by porcine AIDA-I positive <i>E. coli</i> isolates and its receptors. <p>In this study, we demonstrated that AIDA-I adhesin isolated from porcine AIDA-I positive <i>E. coli</i> PD20 and PD58 is an acidic protein consisting of five isoforms. It has a molecular weight (100 kDa) similar to the AIDA-I adhesin expressed by human AIDA-I positive <i>E. coli</i> strain 2787 and has a relatively high amino acid homology (78-87%) with it. Immunodetection of AIDA-I positive <i>E. coli</i> strains using polyclonal anti-AIDA-I antibodies had relatively low sensitivity and specificity, accordingly these tests are unlikely to be used for regular diagnostic detection. <p>Using affinity chromatography, we isolated from porcine intestinal mucus proteins that bind to purified AIDA-I adhesin. These proteins were separated by one- and two-dimensional electrophoresis and subjected to overlay Western blot with purified AIDA-I adhesin and AIDA-I positive <i>E. coli</i> to demonstrate 65 and 120 kDa (p65 and p120) proteins as AIDA-I binding proteins. The identity of p65 was not determined based on LCMS/MS data, whereas p120 was matched to two nuclear proteins (namely, DNA damage binding protein and splicing factor 3b) and one cytoplasmic protein, which is an IgG Fc binding protein. Based on similar amino acid homology, molecular weight, structural similarity to mucin and reported evidence of being secreted by goblet cells into the intestinal lumen, we think that the IgG Fc binding protein is the most likely candidate to serve as a potential receptor in intestinal mucus for AIDA-I adhesin.

Syfte: Redogöra för eventuellt användbara teorier då vi undersöker varför Handelsbanken samt Swedbank har valt att marknadsföra sig som de gör. Metod: För att uppnå vårt syfte har vi valt att ha en kvalitativ utgångspunkt i vår uppsats. Primärdata har samlats in genom personliga intervjuer med anställda på både handelsbanken samt Swedbank. Sekundärdata har samlats in från litteratur och vetenskapliga artiklar. Teoretisk referensram: Den teoretiska referensramen har byggts upp kring relevanta teorier och modeller som har anknytning till profilering av marknadsföringsstrategier. Empiri: Primärdata har samlats in genom personliga intervjuer med kontorschefer på handelsbanken och ansvarig personal för marknadsföringsavdelningen i Swedbank, alla intervjuer har genomförts i Stockholm. Resultat: Det resultat som har nåtts efter att uppsatsen har avslutats är att Handelsbanken marknadsför sig lokalt samt att Swedbank marknadsför sig genom stora kommunikationskanaler för att nå ut till alla konsumenter.

A relatively high percentage of porcine <i>Escherichia coli</i> isolates from cases associated with neonatal and post-weaning diarrhea are positive for the gene encoding the adhesin involved in diffuse adherence I (AIDA-I). This gene and its corresponding protein were first identified and characterized in <i>E. coli</i> strain 2787 isolated from human infantile diarrhea. Little is known about the role of the AIDA-I protein in pathogenesis of porcine enteric disease caused by AIDA-I positive E. coli and the properties of AIDA-I protein expressed by porcine AIDA-I positive <i>E. coli</i> isolates and its receptors. <p>In this study, we demonstrated that AIDA-I adhesin isolated from porcine AIDA-I positive <i>E. coli</i> PD20 and PD58 is an acidic protein consisting of five isoforms. It has a molecular weight (100 kDa) similar to the AIDA-I adhesin expressed by human AIDA-I positive <i>E. coli</i> strain 2787 and has a relatively high amino acid homology (78-87%) with it. Immunodetection of AIDA-I positive <i>E. coli</i> strains using polyclonal anti-AIDA-I antibodies had relatively low sensitivity and specificity, accordingly these tests are unlikely to be used for regular diagnostic detection. <p>Using affinity chromatography, we isolated from porcine intestinal mucus proteins that bind to purified AIDA-I adhesin. These proteins were separated by one- and two-dimensional electrophoresis and subjected to overlay Western blot with purified AIDA-I adhesin and AIDA-I positive <i>E. coli</i> to demonstrate 65 and 120 kDa (p65 and p120) proteins as AIDA-I binding proteins. The identity of p65 was not determined based on LCMS/MS data, whereas p120 was matched to two nuclear proteins (namely, DNA damage binding protein and splicing factor 3b) and one cytoplasmic protein, which is an IgG Fc binding protein. Based on similar amino acid homology, molecular weight, structural similarity to mucin and reported evidence of being secreted by goblet cells into the intestinal lumen, we think that the IgG Fc binding protein is the most likely candidate to serve as a potential receptor in intestinal mucus for AIDA-I adhesin.

The constantly evolving use of social media as a marketing tool is commonly known, and the opportunities for companies to get feedback about how successful their marketing is, have grown just as much. Today's companies take great financial risks when choosing marketing material in the traditional marketing way, but as the social network Instagram is free to use, the biggest risk here is to lose followers, and thus potential customers. This study aims to investigate what consumers actually want to see in the direct marketing they themselves have chosen to receive; what the consumers find most appealing and what makes them feel the greatest desire to buy. For this study seven semi-structured interviews were held with seven women all in the age between 16 and 25. They shared their everyday habits on the social media platform Instagram and were shown 15 Instagram pictures from five major fashion companies. The women expressed their spontaneous feelings when the pictures were shown, just as if the images had been found in their own Instagram feed. The images shown consisted of two, by us, defined messages divided into a five-degree scale, where the scale was ranked accordingly: extremely rational, rational, mixed, emotional and extremely emotional messages. The interviewees also commented on their preferences regarding the content of the pictures, if it was up to their own choosing. After analyzing all interviews, we have come to the conclusion that both the rational and the emotional image categories are preferable to the others. These two stand out from the five-degree scale as both more appealing and creating a greater desire to buy. The extreme ends of each category does not suit marketing on Instagram for different reasons. With this information, we hope to help companies with their marketing strategy on Instagram. This thesis is written in Swedish. / Användningen av sociala medier som marknadsföringsverktyg växer ständigt, och så även möjligheterna för företag att få återkoppling angående hur framgångsrik deras marknadsföring är. Dagens företag tar stora ekonomiska risker när de väljer marknadsföringsmaterial på det traditionella sättet, men när det på sociala mediet Instagram är gratis att skapa ett konto blir den största risken att förlora följare och därmed potentiella kunder. Den här studien undersöker vad konsumenter faktiskt önskar att se i den direktreklam de självmant väljer att följa, vad som mest tilltalar dem och vad som får dem att känna mest köplust. Studien består av sju semistrukturerade intervjuer med kvinnor i åldrarna 16-25 som berättar om sina vanor på det sociala mediet Instagram. De sju kvinnorna visades 15 Instagrambilder från fem stora modeföretag och fick uttrycka sina spontana känslor inför dessa, likt ifall bilderna hade funnits i intervjupersonernas Instagramflöde. Bilderna bestod av två budskap som var uppdelade på en femgradig skala, där skalan rangordnades efter kategorierna extremt rationellt, rationellt, mixat, emotionellt och extremt emotionellt budskap. Intervjupersonerna kommenterade även vilka preferenser de hade angående bildernas innehåll ifall de själva fick välja. Efter att ha analyserat alla intervjuer menar vi att både de rationella och de emotionella bildkategorierna är att föredra, de står ut på den femgradiga skalan som både mer tilltalande och mer skapande av köplust. De extrema kategorierna av båda budskapen passar inte marknadsföring på Instagram av olika skäl. Med den här informationen hoppas vi kan hjälpa företag med deras marknadsföringsstrategi på Instagram.

Thèse numérisée par la Division de la gestion de documents et des archives de l'Université de Montréal.

As produções literárias de Orlanda Amarílis, Ivone Aida e Fátima Bettencourt, sob a forma de conto, constituem o objeto central dessa pesquisa e, a partir do corpus selecionado, buscamos identificar e compreender as opções de construção de personagens e de narração que definem, nas coletâneas Cais-do-Sodré té Salamansa, Vidas vividas e Semear em pó, respectivamente, representações sociais de gênero e características do Bildung (formação da personagem), com base numa hermenêutica do cotidiano cabo-verdiano no arquipélago e na diáspora. No primeiro capítulo, procuramos estabelecer um campo teórico, de largo espectro, que pudesse dar suporte às nossas ilações, para tanto aplicando os vários conceitos que orientaram a nossa visada sobre o objeto da pesquisa, como: entre-lugar, póscolonialismo, hermenêutica do cotidiano, diáspora, Bildungsroman feminino; em seguida passamos a uma incursão no contexto histórico, na biobibliografia das autoras e na conformação das coletâneas, pretendendo situar as obras para o nosso leitor. Iniciamos o processo de análise, logo na primeira parte do segundo capítulo, procedendo a uma investigação sobre os títulos dos contos e sua representação na diegese. Na segunda parte, buscamos investigar com mais detalhe os processos de estruturação das personagens, empreendidos pelas três escritoras, com vista a compreendermos os diversos modos de apreensão e representação do universo cabo-verdiano, especialmente o de vivências femininas de submissão, resistência e emancipação. O terceiro capítulo, que complementa o de panorâmica da construção das personagens, será dedicado a examinar, de forma sucinta, a atuação dos(as) narradores(as), que, em conjunto com as personagens, revelam a mundividência que as escritoras querem registrar na ficção, no intuito de introduzir uma ótica feminina e, por vezes, feminista, ao cânone literário cabo-verdiano, predominantemente masculino. O quarto capítulo foi dedicado a uma sistematização das conclusões. / The central object of this research is based on Orlanda Amarílis, Ivone Aida and Fátima Bettencourts short story production from the selected corpus, we attempted to identify and understand the choice for constructing characters and narration that define in the collections Cais-do-Sodré té Salamansa, Vidas vividas and Semear em pó, respectively, social representations of gender and characteristics of the Bildung (character formation), based on Cape Verdeans hermeneutics of everyday life both in the archipelago and in the diaspora. On the first chapter we tried to establish a broad theoretical field that could support our inferences, thus applying various concepts that oriented our look at the researched object, such as: in-betweenness, post-colonialism, hermeneutics of everyday life, diaspora, the female Bildungsroman. Then we moved to the historical context at the authors bio-bibliography and in the conformation of the collections with the intention to locate the works for our readers. We began the analysis process, on the first part of the second chapter, investigating the titles of the stories and their representation in the diegesis. On the second part, we researched in more detail the processes of structuring characters, undertaken by the three writers, in order to understand the different modes of apprehension and representation of the universe of Cape Verde, especially the experiences of female submission, resistance and emancipation. The third chapter, which complements the overview of the construction of the characters, will be devoted to examine briefly the action of narrators, which together with the characters reveal the worldviews that the writers want to register in fiction in order to introduce a feminine viewpoint and sometimes feminist literary canon to Cape Verde still predominantly masculine. The fourth chapter is devoted to classifying the findings.

Thèse numérisée par la Division de la gestion de documents et des archives de l'Université de Montréal

As produções literárias de Orlanda Amarílis, Ivone Aida e Fátima Bettencourt, sob a forma de conto, constituem o objeto central dessa pesquisa e, a partir do corpus selecionado, buscamos identificar e compreender as opções de construção de personagens e de narração que definem, nas coletâneas Cais-do-Sodré té Salamansa, Vidas vividas e Semear em pó, respectivamente, representações sociais de gênero e características do Bildung (formação da personagem), com base numa hermenêutica do cotidiano cabo-verdiano no arquipélago e na diáspora. No primeiro capítulo, procuramos estabelecer um campo teórico, de largo espectro, que pudesse dar suporte às nossas ilações, para tanto aplicando os vários conceitos que orientaram a nossa visada sobre o objeto da pesquisa, como: entre-lugar, póscolonialismo, hermenêutica do cotidiano, diáspora, Bildungsroman feminino; em seguida passamos a uma incursão no contexto histórico, na biobibliografia das autoras e na conformação das coletâneas, pretendendo situar as obras para o nosso leitor. Iniciamos o processo de análise, logo na primeira parte do segundo capítulo, procedendo a uma investigação sobre os títulos dos contos e sua representação na diegese. Na segunda parte, buscamos investigar com mais detalhe os processos de estruturação das personagens, empreendidos pelas três escritoras, com vista a compreendermos os diversos modos de apreensão e representação do universo cabo-verdiano, especialmente o de vivências femininas de submissão, resistência e emancipação. O terceiro capítulo, que complementa o de panorâmica da construção das personagens, será dedicado a examinar, de forma sucinta, a atuação dos(as) narradores(as), que, em conjunto com as personagens, revelam a mundividência que as escritoras querem registrar na ficção, no intuito de introduzir uma ótica feminina e, por vezes, feminista, ao cânone literário cabo-verdiano, predominantemente masculino. O quarto capítulo foi dedicado a uma sistematização das conclusões. / The central object of this research is based on Orlanda Amarílis, Ivone Aida and Fátima Bettencourts short story production from the selected corpus, we attempted to identify and understand the choice for constructing characters and narration that define in the collections Cais-do-Sodré té Salamansa, Vidas vividas and Semear em pó, respectively, social representations of gender and characteristics of the Bildung (character formation), based on Cape Verdeans hermeneutics of everyday life both in the archipelago and in the diaspora. On the first chapter we tried to establish a broad theoretical field that could support our inferences, thus applying various concepts that oriented our look at the researched object, such as: in-betweenness, post-colonialism, hermeneutics of everyday life, diaspora, the female Bildungsroman. Then we moved to the historical context at the authors bio-bibliography and in the conformation of the collections with the intention to locate the works for our readers. We began the analysis process, on the first part of the second chapter, investigating the titles of the stories and their representation in the diegesis. On the second part, we researched in more detail the processes of structuring characters, undertaken by the three writers, in order to understand the different modes of apprehension and representation of the universe of Cape Verde, especially the experiences of female submission, resistance and emancipation. The third chapter, which complements the overview of the construction of the characters, will be devoted to examine briefly the action of narrators, which together with the characters reveal the worldviews that the writers want to register in fiction in order to introduce a feminine viewpoint and sometimes feminist literary canon to Cape Verde still predominantly masculine. The fourth chapter is devoted to classifying the findings.