Laboratory directed evolution have enabled the discovery of numerous functional natural and synthetic macromolecules with tailor-made functions. However, approaches that use enzymes to effect the crucial translation from an information carrier molecule such as DNA or RNA to synthetic polymers are limited to producing close analogs of nucleic acids, either due to a strict requirement to hybridize with a nucleic acid template or as a consequence of the limited substrate scope of polymerase enzymes. / Chemistry and Chemical Biology

abstract: Protein affinity reagents have aptly gained profound importance as capture reagents and drugs in basic research, biotechnology, diagnostics and therapeutics. However, due to the cost, labor and time associated with production of antibodies focus has recently changed towards potential of peptides to act as protein affinity reagents. Affinity peptides are easy to work with, non-immunogenic, cost effective and amenable to scale up. Even though researchers have developed several affinity peptides, we are far from compiling library of peptides that encompasses entire human proteome. My thesis describes high throughput pipeline that can be used to develop and characterize affinity peptides that bind several discrete sites on target proteins. Chapter 2 describes optimization of cell-free protein expression using commercially available translation systems and well-known leader sequences. Presence of internal ribosome entry site upstream of coding region allows maximal expression in HeLa cell lysate whereas translation enhancing elements are best suited for expression in rabbit reticulocyte lysate and wheat germ extract. Use of optimal vector and cell lysate combination ensures maximum protein expression of DNA libraries. Chapter 3 describes mRNA display selection methodology for developing affinity peptides for target proteins using large diversity DNA libraries. I demonstrate that mild denaturant is not sufficient to increase selection pressure for up to three rounds of selection and increasing number of selection rounds increases probability of finding affinity peptide s. These studies enhance fundamental understanding of mRNA display and pave the way for future optimizations to accelerate convergence of in vitro selections. Chapter 4 describes a high throughput double membrane dot blot system to rapidly screen, identify and characterize affinity peptides obtained from selection output. I used dot blot to screen potential affinity peptides from large diversity of previously ii uncharacterized mRNA display selection output. Further characterization of potential peptides allowed determination of several high affinity peptides from having Kd range 150- 450 nM. Double membrane dot blot is automation amenable, easy and affordable solution for analyzing selection output and characterizing peptides without ne ed for much instrumentation. Together these projects serve as guideline for evolution of cost effective high throughput pipeline for identification and characterization of affinity peptides. / Dissertation/Thesis / Masters Thesis Biochemistry 2014

Cultural adaptation and free translation strategies are more common to use within the field of video game localization although they are still not recommended too veruse in translation studies. Free translation strategies are commonly used invideo game localization to make video games ready for a targeted audience. As free translation is mostly target-oriented in video game localization, the original source text often loses its meaning because of culturally adapting and modifying the text to fit to the target culture. This study aims to analyze 10 chosen cutscenes of the video game Lost Judgement, to find how frequently cultural adaptation and free translation strategies were used for pragmatic (proper names, cultural termsand expressions, allusion) and interlingual (regional dialect and slang) translation problems and how these problems were solved. The result and analysis of the study demonstrate examples of these translation problems within chosen cutscenes. The examples prove that depending on translation problems, both free translation and cultural adaptation can be used in combination with a literal word-for-word translation strategy. It also indicates that the video localization of Lost Judgement implied to use both domestication and foreignization approaches to achieve a higher satisfaction for the players of the game who can choose their favorite version. Another important factor was that the localization of the videogame focused on a functional approach whose purpose was to keep loyalty to the original text, the source text.

In this thesis, medieval free translation is explored as a text-producing practice as it appears in Alexanders saga, a 13th century Old Norse translation of the medieval Latin epic Alexandreis. The practice is investigated through analyses of (1) the rendering of the source text and (2) the translator’s role in making the target text. The rendering is analyzed through a systematic comparison between source and target text using a method of analysis based on systemic functional linguistics (SFL). Contrary to what was assumed previously, the rendering pro­ves to be consistent in the text, but a surprising result is that the rendering in chapters 2–4 and in chapters 1 and 6–10 respectively represent two significantly dif­ferent patterns, the former being closer to the source text than the latter, pre­sumably due to two different translators. The investigation further confirms an observation in previous research on Old Norse free translation that the rendering of parts in direct speech are closer to the source than that of narrative and descriptive discourse. The rendering is closest where the translator indicates that he is quoting the author of the source text. These patterns are found in both groups of chapters, and as they are confirmed in other Old Norse translations, they might be interpreted as a translation norm. The conceptions of translation are further investigated by examining what kind of text-producing role the translator assumes. It is claimed that, despite the freedom in free ren­dering, the translator assumes the role of intermediary between the source text and the receivers of the target text rather than the role of independent text pro­ducer. From an analysis of the translator’s metatextual additions, it seems as though this is also what the translator assumes the receivers of the text expect him to do. The results indicate the presence of certain conceptions of how translation was to be carried out in West Nordic society. The ”free” translation strategies did not mean freedom from or obliviousness to translation norms, but rather re­late to a specific text-producing practice.

Made available in DSpace on 2016-04-29T14:17:57Z (GMT). No. of bitstreams: 1 Cristiane Andrade Garcia.pdf: 2111056 bytes, checksum: d37b3c38bbaddcd5f6d9ce8f1fac6eb0 (MD5) Previous issue date: 2009-10-23 / This present research was conducted on the current context of implementing AMAs Assistência Médica Ambulatorial (Ambulatory Medical Assistance) in the city of São Paulo, specifically that located in Vila Nova Jaguaré zone. This AMA is a partnership between Prefeitura Muncipal de São Paulo and Fundação Faculdade de Medicina. It is a study on this service and the insertion of Social Service on this scenario. The field research was developed at AMA-Jaguaré. This study plunged into concepts related to health care, the relationship between private organizations and public assistance, Social Service practice on health care and the professional competence, the work management in a multi-professional team. The methodology has embraced the bibliographic research on the main themes, besides the documental research (laws, manuals and reports), participative investigation and the quantitative/qualitative field research. Concerning the qualitative one, the interview and the semi-structured interview were used in users, employees and professional s approach methodological procedures. The results which were obtained have shown that the service rendered by AMA may have created a new demand within the assistance net, because it showed a search that we can consider as high. We can affirm that there was not a proper projection concerning the assistance demand volume that would be created, considering the limitation of physical space and lack of professionals. Besides, it was evidenced a trend among the users concerning the need of indispensable equipment, such as XR device, for more specific health diagnosis, resulting in a demand of such arrangement. Simultaneously, it was also raised the importance of a higher attention on the reference process, by sending the cases to specialties. In terms of a positive assessment of AMA, it was made explicit the work role from Social Service. Other professionals have recognized it as a differential on user welcome, listening, mediation and attention / A presente pesquisa contextualiza-se no momento atual, de implementação das AMAs - Assistência Médica Ambulatorial na cidade de São Paulo, especificamente daquela situada no bairro de Vila Nova Jaguaré, como parte de uma parceira entre a Prefeitura Municipal de São Paulo e a Fundação Faculdade de Medicina. Constitui um estudo sobre esse serviço e a inserção do Serviço Social nesse contexto. A pesquisa de campo foi desenvolvida na AMA Jaguaré, sendo nela aprofundados alguns conceitos relativos à saúde, à relação de organizações privadas com atendimento público, à prática do Serviço Social na saúde, à competência profissional e à gestão do trabalho em equipe multiprofissional. A metodologia abrangeu pesquisa bibliográfica sobre os temas centrais, além de pesquisa documental (leis, manuais e relatórios), observação participante e pesquisa de campo quanti-qualitativa. Quanto a esta última, foram utilizados como procedimentos metodológicos de abordagem de usuários, funcionários e profissionais, questionário e entrevista semiestruturada. A pesquisa foi norteada por algumas questões básicas, levando-se em conta principalmente a forma como foi pensada em São Paulo a questão das Organizações Sociais no âmbito da saúde, na relação público / privado. Os resultados obtidos evidenciaram que o serviço prestado pela AMA pode ter criado uma nova demanda dentro da rede de atendimento, ao ter recebido uma procura que deve ser considerada alta. Pode-se afirmar que não foi feita uma projeção adequada do volume da demanda de atendimento que seria criada, tanto se considerando a limitação do espaço físico como a insuficiência de profissionais. Além disso, constatou-se entre os usuários um consenso sobre a necessidade de equipamentos indispensáveis, tais como aparelho de RX para diagnósticos de saúde mais específicos, tendo sido reivindicada essa providência. Foi levantada também, paralelamente, a importância de maior atenção ao processo de referência, com a realização de encaminhamentos para especialidades. Em termos de avaliação positiva da AMA, explicitouse o papel do trabalho desenvolvido pelo Serviço Social, que foi reconhecido pelos outros profissionais como um diferencial na abordagem, na escuta, na mediação e no acolhimento do usuário

Synthesizing proteins for use in therapeutics is restrained by, in part, contaminants in in vivo expression systems and limited production capacity of in vitro systems. Cell free expression (CFE) systems have emerged as a potential alternative for protein expression because of the inherently lower contents of contaminants, and their flexible modular design that allows the addition of factors that aid in synthesis of complex products. Here, we investigate and establish an in-house Escherichia coli-based cell free protein synthesis (CFPS) system, explore different CFPS commercial kits, develop assays to test performance of these systems and identify potential rules that dictate expression levels. Using CFE, we were able to test different vectors and conditions of system, as well as scale-up protein synthesis reactions. In conclusion, this work shows that CFPS is a functional and easy-to-use platform and can potentially meet the requirements for the synthesis of therapeutics.