The type II secretion system (T2SS) is widespread in Gram-negative bacteria that cause disease in animals and plants. In human and animal pathogens toxins are secreted (e.g. cholera toxin) and in plant pathogens lytic enzymes that breakdown the plant cell wall are exported in to the extracellular milieu (e.g. pectate lyase). Structurally the T2SS comprises at least 11 core proteins that form three major subassemblies spanning the inner-membrane, periplasmic space and outer-membrane: (i) the inner-membrane platform and associated cytoplasmic ATPase (E); (ii) the pseudopilus, which consists of five pseudopilins, G to K; and (iii) a large, pore-forming outer-membrane complex secretin D. The inner-membrane platform comprises three single transmembrane helix proteins, and one three transmembrane helix protein, OutF. The evidence from cryo-electron microscopy on the related type IVa pilus machine (T4PS) places the protein corresponding to OutF at the centre of this platform. This platform is responsible for assembling the pilus and for communicating between the periplasm and the cytoplasmic ATPase. To date, no high-resolution structure of a full-length OutF/PilC family protein is available. A low-resolution electron microscopy reconstruction of isolated PilG (PilC ortholog from Neisseria meningitides T4PS) showed a tetrameric two lobed structure. Here I report the results of studying the structure of the inner-membrane protein OutF from Dickeya dadantii and the complete inner-membrane platform comprising 9 proteins: OutEFGHIJKLM. This work involved cloning the corresponding operon, purifying the proteins, and using crystallography and electron microscopy. Key results reported here are the crystal structure of the first cytoplasmic domain of Dickeya dadantii, OutF65-172 and a preliminary three-dimensional model of the Dickeya dadantii inner-membrane platform. This model, and higher-resolution models to come, will provide valuable information about the oligomeric state, and arrangement of the inner-membrane proteins. These studies will help us to understand how the type II secretion system works.

No description available.

This thesis examines the attitudes of British people to the aerial bombardment of German cities during the Second World War, with particular attention given to those who challenged the nature of the campaign. I use contemporary sources with a strong emphasis on qualitative data to develop a picture of attitudes at the time and situate the roots of the significant post-war controversy within these contemporary attitudes. The thesis offers a more sustained and textured account of anti-bombing sentiment than other historiographical works. An introductory chapter charts the development of aerial bombing in the early years of the twentieth century. The extent to which Britain engaged with aerial bombardment, and how it was understood by people in Britain, are addressed here. Three case studies – each focusing on a different raid on a German city – are then used to address how attitudes to the bomber offensive were shaped at different stages of the war. The first is the December 1940 attack on Mannheim. This took place during the Blitz on British cities, a factor which has implications for the nature of responses at this time. The question of reprisals is important here. I show how the desire for reprisals was far from universal, yet it was overstated in the press and by Prime Minister Winston Churchill. The second case study addresses the series of heavy attacks on Hamburg in July and August 1943. This followed the decision, taken the previous year, to officially adopt a policy of area bombing. This chapter shows how the Archbishop of Canterbury's support for the campaign stifled voices of protest at this time. The final case study considers the raids on Dresden in February 1945. Churchill's response is addressed in this chapter and contrasted with the immediate concerns raised in the press and in private diaries.

This thesis is a detailed analysis of British foreign policy between 3 September 1939 and 10 May 1940. It concentrates on policy towards the Far East, Italy, the Soviet Union, the Balkans, and Scandinavia. These areas represented the biggest challenges to British policy following the outbreak of war with Germany: Japan and Italy, whilst nominal allies of Germany, had opted to stay out of the war, the Soviet Union appeared to be acting in collaboration with Germany but was not at war with Britain, and the Balkans and Scandinavia were the most likely theatres of war if the conflict were to spread. Lack of resources dictated that British efforts be directed towards minimising military activity and containing the conflict, whilst putting economic pressure on Germany’s ability to fight. Potential allies of Germany had to be dissuaded from entering the war and prevented from helping Germany economically. Potential theatres of war had to be kept neutral unless or until an extension of hostilities would be in Britain’s interests. The contradictions and conflicts of interest created by these policies posed serious problems, and it is the British attempts to solve these problems which form the focus of this study.

Abstract not available

<p><strong>Problem: </strong>Hur har implementeringen av det nya regelverket Basel II påverkat en liten respektive en stor bank? Vilken metod använder en liten respektive en stor bank vid beräkning av risker enligt den första pelaren och är storleken på bank avgörande för val av metod? Vilka brister har Basel II och hur kan regelverket förbättras?</p><p><strong>Syfte: </strong>Denna studie syftar till att beskriva och analysera hur Basel II påverkat en liten respektive en stor bank, dessutom påvisa om det finns skillnader i val av metod vid beräkning av risker enligt den första pelaren. Syftet är även att belysa Basel II´s brister, samt föreslå möjliga förbättringar.</p><p><strong>Metod: </strong>Vi har valt en kvalitativ undersökningsmetodik för insamling av empirisk data. Undersökningen bestod av intervjuer med en liten respektive en stor bank, samt Finansinspektionen. Även böcker, rapporter och tidigare forskning har legat till grund för vår studie.</p><p><strong>Slutsats: </strong>Bankerna i den här studien anser att implementeringen av Basel II varit resurskrävande, både kostnads- och kompetensmässigt. Nyttan har dock övervägts av kostnaderna då både Sparbanken i Enköping och storbanken ökat sin riskhanteringsförmåga. Val av metod skiljer sig enbart vid beräkning av kreditrisker. Storbanken använder den grundläggande IRK-metoden och har därför haft fördelen att få en kapitallättnad. Brister i det nuvarande regelverket är att kapitalkravet blir volatilt över tiden och att likviditetsrisken inte fått ett tillräckligt stort utrymme, vilket kan förbättras i ett framtida regelverk.</p>

Long-term stimuli of many systems leads to decreased cellular responsiveness, or desensitization. We characterized the desensitization of angiotensin II (Ang 11)-mediated inositol phospholipid (IP) hydrolysis in cultured human aortic smooth muscle cells (HASMC). Although it has been suggested that the desensitization induced by long-term Mg II exposure may result partially from down-regulation of Ang II receptor, this is not sufficient to explain fully desensitization in many systems. Post-receptor desensitization of IP hydrolysis may also result from phosphorylation or changes in protein levels of the effector enzyme, PLC-β. We identified the major PLC-β isoenzymes expressed by HASMC as PLC-β1 and PLC-β3. Ang II pretreatment reduced IP accumulation induced by Ang II (1μM) in a time-dependent manner. Phorbol ester-12-myristrate-13-acetate (PMA), a protein kinase C (PKC) activator, also reduced Ang II-stimulated IP accumulation. These results suggest that PKC activation may negatively regulate Ang II-stimulated IP signaling in HASMC, similar to rat cells. In addition, PKC also reduced IP accumulation stimulated by A1F₄⁻, directly activating the G protein. It suggests that the majority of PKC-induced desensitization of Ang II-stimulated IP signaling occurs downstream of the Ang II receptor in HASMC. However, both PLC-β1 and PLC-β3, expected candidates for PKC phosphorylation, were phosphorylated independently of PKC activation or inhibition, indicating that PKC might not be involved in direct phosphorylation of PLC-β1 and PLC-β3. Furthermore, PLC-β1, but not PLC-β3, was highly phosphorylated under basal conditions, suggesting that PLC-β1 and PLC-β3 may play different roles in IP signaling in HASMC. / Graduation date: 2003

Originally issued as Thesis--Universität Bonn, 2003/2004. / Includes bibliographical references (p. 148-162).

Problem: Hur har implementeringen av det nya regelverket Basel II påverkat en liten respektive en stor bank? Vilken metod använder en liten respektive en stor bank vid beräkning av risker enligt den första pelaren och är storleken på bank avgörande för val av metod? Vilka brister har Basel II och hur kan regelverket förbättras? Syfte: Denna studie syftar till att beskriva och analysera hur Basel II påverkat en liten respektive en stor bank, dessutom påvisa om det finns skillnader i val av metod vid beräkning av risker enligt den första pelaren. Syftet är även att belysa Basel II´s brister, samt föreslå möjliga förbättringar. Metod: Vi har valt en kvalitativ undersökningsmetodik för insamling av empirisk data. Undersökningen bestod av intervjuer med en liten respektive en stor bank, samt Finansinspektionen. Även böcker, rapporter och tidigare forskning har legat till grund för vår studie. Slutsats: Bankerna i den här studien anser att implementeringen av Basel II varit resurskrävande, både kostnads- och kompetensmässigt. Nyttan har dock övervägts av kostnaderna då både Sparbanken i Enköping och storbanken ökat sin riskhanteringsförmåga. Val av metod skiljer sig enbart vid beräkning av kreditrisker. Storbanken använder den grundläggande IRK-metoden och har därför haft fördelen att få en kapitallättnad. Brister i det nuvarande regelverket är att kapitalkravet blir volatilt över tiden och att likviditetsrisken inte fått ett tillräckligt stort utrymme, vilket kan förbättras i ett framtida regelverk.

<i>Aeromonas hydrophila</i> uses the type II secretion system to transport protein toxins across the outer membrane. The trans-envelope system is comprised of more than ten proteins, including ExeA and ExeB, which form a complex in the inner membrane and are required for assembly of the ExeD secretion channel multimer, called the secretin, into the outer membrane. A putative peptidoglycan binding domain (Pfam protein families database number PF01471) is present in the periplasmic region of ExeA (pExeA), leading to the hypothesis that ExeA generates gaps in peptidoglycan, a barrier for trans-envelope transport and apparatus assembly, to allow ExeD to assemble into the outer membrane.<p> In this study, interactions between ExeA and peptidoglycan were examined both <i>in vivo</i> and <i>in vitro</i>. Wild type ExeA, but not the mutants containing substitution mutations of three highly conserved amino acid residues in the putative peptidoglycan binding domain, was cross-linked to peptidoglycan in vivo with DTSSP. Furthermore, the presence of wild type ExeA was also required for co-crosslinking of ExeB and ExeC to peptidoglycan. <i>In vitro</i> cosedimentation revealed that purified pExeA was able to bind to highly purified peptidoglycan. The protein assembled into large multimers in the presence of peptidoglycan fragments, as shown in cross-linking and co-gel filtration experiments. The requirement of peptidoglycan for multimerization was abrogated when the protein was incubated at temperatures above 25 °C. Two pExeA constructs, which disrupted the putative peptidoglycan binding domain, greatly reduced the cosedimentation, accompanied by decreased multimerization in the presence of peptidoglycan fragments. These results provide evidence that the putative peptidoglycan binding domain of ExeA is involved in physical contact with peptidoglycan. The interactions cause ExeA to multimerize, possibly forming a ring-like structure on the peptidoglycan, to generate a gap large enough to accommodate the secretion apparatus and/or to form an assembly scaffold.<p> The putative peptidoglycan binding domain of ExeA was also analyzed by comparing its amino acid sequence with that of other homologues. The highly conserved amino acid residues were found to cluster at one pocket on the surface in the crystal structure of hydrolase metallo (Zn) DD-peptidase that also contains this domain. We propose that this pocket is the binding site for the peptidoglycan ligand.