Cytochromes P450 (P450s) are a superfamily of heme-thiolate monooxygenases. They catalyse a wide variety of reactions on a vast number of substrates and are of particular interest for biocatalyst development due to their ability to oxidise non-activated C-H bonds. Fusion of a P450 to a suitable redox partner protein produces a catalytically self-sufficient enzyme and removes the need to produce electron transfer proteins separately. The well-studied bacterial protein P450cam (Pseudomonas putida) has been fused to the reductase (RhFRed) from the natural fusion protein P450-RhF (Rhodococcus sp.). The P450cam-RhFRed system catalyses the oxidation of camphor and several non-natural substrates and served as the basis for P450cam re-engineering in this current project, with the aim of expanding the substrate scope towards a more mammalian-like activity. The P450cam active site was partitioned into seven paired amino acids and each pair randomised in turn to generate seven sub-libraries of P450cam variants. These were screened for activity using a specially developed colony screen for detection of the blue pigment indigo. In total 94 new variants were identified and then pooled for secondary screening on a number of new substrates, identifying potentially novel activities within the ‘indigo positive’ population. In a separate ‘chimeragenesis’ approach substrate recognition sites (SRSs) within P450cam were targeted for exchange with equivalent portions from a number of human P450s. The B’ helix and F-G loop regions from CYPs 1A2, 2C8, 2D6 and 3A4 were grafted onto the P450cam structure and several of the B’ helix swaps were produced as soluble proteins. The P450cam-2C8-B’-RhFRed chimera gave a Soret peak at 420 nm in the Fe(II)-CO state although an additional substitution next to the proximal cysteine appeared to restore a P450-like state. SRS-exchange therefore offered some insight into structural modularity in P450s, providing a basis for further biocatalyst development.

Diabetes is a disease with a deleterious pathology that currently impacts 4.5 million individuals within the United States. This study examined the ability of a specific neuropsychological battery to identify and classify dementia type, investigated the impact of diabetes on cognition and analyzed the ability of the memory measures of the 7 Minute Screen (7MS) and the Rey-Osterrieth Recall to correctly categorize dementia type when not used in combination with a full battery. The battery in addition to exhaustive patient history, medical chart review and pertinent tests were used in initial diagnosis. Results indicated the battery was sufficient in the identification and classification of dementia type. Within the sample, diabetes did not appear to significantly impact overall battery results whereby only two measures were minimally affected by diabetes. Finally, the memory measures of the 7MS and the Rey-Osterrieth Recall were sufficient to predict membership into the Alzheimer's (AD) and vascular dementia (VD) groups with 86.4% accuracy. The classification percentage dropped to 68.3% with addition of the mild cognitive impairment category. The full battery correctly classified AD and VD dementia 87.5% and appeared to be the most robust.

Although many theorists have speculated that chronic pain may be linked to some sort of central neuropsychological integration deficit, a review of the current literature reveals no empirical support for this theory. This study attempts to assess the severity, if any, of neuropsychological deficits in chronic pain subjects by using a neuropsychological screen developed at the University of North Texas. Also, presented are studies of correlations between the UNT Neuropsych-screen and the MMPI. the Dallas Pain Questionnaire (DPQ), the Dallas Pain Drawing CDPD), and time since injury in order to assess any possible relationships. The subjects in this study consist of 100 volunteers. Of these subjects, 74 were patients of the Spinal and Chronic Pain Center at Medical Arts Hospital in Dallas, Texas and represented the clinical population. The remaining 26 subjects were staff volunteers from the hospital . The results of the study indicate significant differences between chronic pain subjects and non-pain subjects across many areas of neuropsychological functioning, as well as other significant correlations among many of the variables. The implications of this study are elaborated upon, in the discussion section, in detail along with limitations and future research directions.

Searching, accessing, reading, and storing academic literature is increasingly done with the help of digital devices. This master’s thesis presents the results of a qualitative study on researchers’ academic reading practices, including the surrounding practices of searching for and storing the literature. The aim of this thesis is to gain a deeper understanding of researchers' use of academic literature on paper and on screens. Semi-structured interviews were conducted with ten researchers within four different research areas from a university college in Norway. The theoretical framework used to discuss the results is practice theory, with the concepts materiality and affordance. Key findings are that there is not always concordance between what the researchers say they prefer and what they do, as they have a pragmatic approach. The respondents see pros and cons of both formats, and they use both, depending on the situation and what their aim is. The surrounding practices are often done digitally, while deep reading is preferred to do on paper, by the majority. The first reading of a text, which is browsing through it to see whether the text is relevant or not, is always done on screen, while most of the respondents prefer to print out articles when they are to be read thoroughly, as they value the physical properties of paper. However, the respondents of this study are more positive to screen reading than what is seen in previous research, and many of them wish to improve their skills when it comes to e-reading.Keywords: Academic reading, researchers, digital reading, reading practices,

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Many of the cutting-edge innovations on the CRISPR/Cas9 system involve the use of complex sgRNA libraries. For example, such libraries have recently been used in functional genomics applications to screen for specific genes in both cell lines and living organisms, as well as in subcellular imaging to fluorescently label chromatin in living cells. However, chemically synthesizing customized libraries can be cost prohibitive, requires precise genomic information, and can takes several weeks. We developed a rapid and efficient enzymatic method to generate sgRNA libraries from arbitrary DNA substrates, significantly reducing the cost of library generation. A type IIS restriction enzyme binding site was incorporated directly into the sgRNA scaffold sequence without affecting the catalytic properties of the Cas9 complex, resulting in a simple method that generates high fidelity sgRNA libraries. Additionally, the library is constructed on the surface of streptavidin coated magnetic beads and a strand displacing polymerase is used to elute DNA from the beads, reducing the loss of material and simplifying purification between steps. We used this method to generate two sgRNA libraries. The first targets the entire E. coli genome, and the second targets genes expressed in the developing zebrafish heart. This second library will be used in a forward genetic screen in zebrafish to identify genes underlying heart defects. These complex sgRNA libraries show the utility of the method in generating libraries from DNA from any species, ranging from prokaryotes like bacteria, to eukaryotes including plants and animals.

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Brand-lurking Phenomenon on Social Media Abstract The key measures to assess consumer-brand engagement on social media have mainly been visible interactions with media content, including 'likes', comments and shares. However, as research into consumers' online behaviour demonstrates, only a minority of users in fact participate 'visibly'. On social media, the vast majority of users are passive participants, so called 'lurkers'. This thesis explores consumers' publicly invisible behaviours on social media and introduces an original methodological approach to exploring the otherwise invisible behavioural patterns on social media, which builds on ethnographic principles and combines digital methods with qualitative methods. Drawing on a data set of 134 hours of screen recordings obtained through ad hoc tracking devices installed on PCs and smartphones of 15 young adults from seven European countries recruited from the "Generation Z" and 15 in- depth interviews conducted with the same participants, the study proposes the following contributions. First, the thesis illustrates a research protocol to explore consumers' lurking behaviours in relation to branded content as they occur within the young people's natural social media settings. Second, it introduces the concept of invisible engagement, defined as an...

This study presents various methods of measuring ozone and consolidates current state of knowledge in the field of disposable printed ozone dosimeters. The paper also identifies prospective dyes for preparation of ozone dosimeters available on the market. Information gathered from literature review was used to prepare dosimeters based on two different dyes. These were calibrated for screen printing technology and their sensitivity to ozone was optimized to achieve visual detectability. Prepared samples were observed and studied while being exposed to ozone. Then, long term mechanical and chemical properties of these were tested in order to identify appropriate conditions for storage.

This thesis deals with issues of used hot plates and thick-film technology. The theoretical part of diploma thesis contains a theoretical analysis and description of corresponding technology and usage of materials. The practical part of the thesis focuses on the design of the resulting device, which forms design of the test and optimized hot element, the initial design of the device and the design of models for temperature simulations. Then follows description of the practical realization of the motifs and testing of the created thick-film structure on a ceramic substrate. In the last part are shown simulations of heat transfer in the proposed model of the device.