<p>This thesis addresses different aspects of the question about accuracy of protein synthesis: i) the mechanism of tRNA selection during translation ii) study of ribosomal mutations that affect accuracy and iii) the choice of aminoacyl-tRNA isoacceptors on synonymous codons.</p><p>By measuring the codon reading efficiencies of cognate and near-cognate ternary complexes we demonstrate that in optimal physiological conditions accuracy of substrate selection is much higher than previously reported; that during translation the ribosomal A site is not blocked by unspecific binding of the non-cognate tRNAs which would inhibit the speed of protein synthesis. Our results suggest that there is an asymmetry between initial selection and proofreading step concerning the wobble position, and that binding of non-cognate substrate does not induce GTP hydrolysis on the ribosome.</p><p>The knowledge obtained from the ribosomal mutant strains can be used to explain the general relation between the structure of the ribosome and the mechanism of codon recognition, as well as the streptomycin resistance or dependence phenomenon.</p><p>Our work showed experimentally that the probability for binding certain tRNA to the A site of the ribosome is not based on the simple codon-anticodon base pair matching. In the living cell the availability of cognate tRNAs versus the demand for them (the frequency of codon usage) is finely balanced to ensure critical protein synthesis in stress conditions. We have also discovered a new codon assignment for a specific tRNALeu isoacceptor and detected a base modification in its anticodon, which has not been previously observed. The motivation for the later findings comes from a system biology modeling and the results are an example of an interdisciplinary collaboration.</p>

This thesis addresses different aspects of the question about accuracy of protein synthesis: i) the mechanism of tRNA selection during translation ii) study of ribosomal mutations that affect accuracy and iii) the choice of aminoacyl-tRNA isoacceptors on synonymous codons. By measuring the codon reading efficiencies of cognate and near-cognate ternary complexes we demonstrate that in optimal physiological conditions accuracy of substrate selection is much higher than previously reported; that during translation the ribosomal A site is not blocked by unspecific binding of the non-cognate tRNAs which would inhibit the speed of protein synthesis. Our results suggest that there is an asymmetry between initial selection and proofreading step concerning the wobble position, and that binding of non-cognate substrate does not induce GTP hydrolysis on the ribosome. The knowledge obtained from the ribosomal mutant strains can be used to explain the general relation between the structure of the ribosome and the mechanism of codon recognition, as well as the streptomycin resistance or dependence phenomenon. Our work showed experimentally that the probability for binding certain tRNA to the A site of the ribosome is not based on the simple codon-anticodon base pair matching. In the living cell the availability of cognate tRNAs versus the demand for them (the frequency of codon usage) is finely balanced to ensure critical protein synthesis in stress conditions. We have also discovered a new codon assignment for a specific tRNALeu isoacceptor and detected a base modification in its anticodon, which has not been previously observed. The motivation for the later findings comes from a system biology modeling and the results are an example of an interdisciplinary collaboration.

Thesis (Ph. D.)--Illinois State University, 2006. / Title from title page screen, viewed on June 8, 2007. Dissertation Committee: Janice Neuleib (chair), Ronald Fortune, Bob Broad. Includes bibliographical references (leaves 270-280) and abstract. Also available in print.

Abstract Replicative DNA polymerases (pols) synthesize chromosomal DNA with high accuracy and speed during cell division. In eukaryotes the process involves three family B pols (α, δ, ε), whereas in Archaea, two types of pols, families B and D, are involved. In this study the B-subunits of replicative pols were analysed at the DNA, RNA and protein levels. By cloning the cDNAs for the B-subunits of human and mouse pol ε we were able to show that the encoded proteins are not only homologous to budding yeast pol ε, but also to the second largest subunit of pol α. Later studies have revealed that the B-subunits are conserved from Archaea to human, and also that they belong to the large calcineurin-like phosphoesterase superfamily consisting of a wide variety of hydrolases. At the mRNA level, the expression of the human pol ε B-subunit was strongly dependent on cell proliferation as has been observed for the A-subunit of pol ε and also for other eukaryotic replicative pols. By analysing the promoter of the POLE2 gene encoding the human pol ε B-subunit we show that the gene is regulated by two E2F-pocket protein complexes associated with the Sp1 and NF-1 transcription factors. Comparison of the promoters of the human pol ε and the pol α B-subunit indicates that the genes for the B-subunits may be generally regulated through E2F-complexes whereas adjustment of the basal activity may be achieved by distinct transcription factors. To clarify the function of the B-subunits, we screened through the expression of 13 different recombinant B-subunits. Although they were mainly expressed as insoluble proteins in E. coli, we were able to optimize the expression and purification for the B-subunit (DP1) of Methanococcus jannaschii pol D (MjaDP1). We show that MjaDP1 alone was a manganese dependent 3'-5' exonuclease with a preference for mispaired nucleotides and single-stranded DNA, suggesting that MjaDP1 functions as the proofreader of archaeal pol D. So far, pol D is the only pol family utilising an enzyme of the calcineurin-like phosphoesterase superfamily as a proofreader.

Este trabalho estuda o campo da edição de texto na produção editorial de livros no Brasil. O objetivo é apontar algumas distinções necessárias às pesquisas acadêmicas sobre edição de textos, e importantes para a formação de pesquisadores e profissionais. A proposta centra-se nas principais etapas (edição de texto, preparação de originais e revisão de provas) que compõem o processo por que passa um texto a ser transformado em livro. Parte-se de análise dos manuais de editoração e de pesquisas acadêmicas que tangenciam o tema. E articula-se um diálogo entre alguns trabalhos acadêmicos das áreas de psicolinguística e psicologia cognitiva sobre leitura em suas relações possíveis com os estudos em editoração e a experiência empírica dos editores de texto. Contribui-se, assim, para o aprimoramento dos conhecimentos sobre a edição de texto, entre eles: as etapas, os objetivos principais e secundários, as estratégias, as tarefas e um conjunto de elementos necessários para que um profissional atue como editor de texto consciente de sua intervenção. / It is the purpose of this thesis to study the field of text-editing in the Brazilian book publishing industry. Its main aim is to point out a few distinctions deemed necessary not only to the academic research but also to the formation of researchers and professionals in this particular field of study and work. In order to achieve this goal, the main phases the text goes through on its way to becoming a book -- text-editing, copy-editing and proofreading -- are here taken into account and characterized. Examining the available manuals and academic papers on the book editing field is the starting point of this research. It then goes on to promoting a dialogue between the book editing field and other areas of expertise, such as psycholinguistics and cognitive psychology, that also deal with the process of reading. The possible relationships this dialogue reveals and the actual praxis of text editing are then analyzed. Contributing to a better knowledge of what text-editing actually involves is therefore a primary aim of this thesis. It intends to shed some light on the specific phases, primary and secondary goals and strategies text-editing comprises, as well as on a series of elements needed not only for the praxis of text-editing itself but also for a more conscientious practice of this activity.

We have developed a novel, sensitive and less time-consuming method to detect activity of the SecA-dependent protein-conducting channels. Nanogram levels of E. coli inverted membrane vesicles were injected into Xenopus oocytes, and ionic currents were recorded using the two-electrode voltage clamp. Currents were observed only in the presence of E. coli SecA in conjunction with E. coli membranes. The observed currents showed outward rectification in the presence of KCl as permeable ions and were significantly enhanced by coinjection with the precursor protein, proOmpA, or active LamB signal peptide. Channel activity was blockable with sodium azide or adenylyl 5’-(β, γ-methylene)-diphosphonate, a non-hydrolyzable ATP analog, both of which are known to inhibit SecA protein activity. Channel activity was also stimulated by oocyte endogenous precursor proteins, which could be inhibited by puromycin. In the presence of puromycin, exogenous proOmpA or LamB signal peptides, but not defective signal peptides, stimulated the ionic currents. We also measured SecA-dependent currents with membranes depleted of SecYEG. Wild-type LamB signal peptides, or precursor proteins stimulated ionic currents following a co-injection of SecYEG¯ membranes with puromycin. Excess exogenous SecA stimulated ionic currents through SecYEG¯ membranes. Similar activities of added SecA were observed with reconstituted membranes depleted of SecYEG. Currents through such SecYEG-depleted membranes were also stimulated by addition of defective LamB signal peptides and unfolded mature PhoA protein. In contrast, currents produced by the membranes containing wild-type SecYEG were not so stimulated, but ionic currents were stimulated through mutant strains, similar to PrlA (SecY) suppressors, e.g. PrlA4, or PrlA665 membranes, suggesting that the proofreading function of SecY was bypassed in these membranes. We have observed that azide can inhibit ionic currents when E. coli wild-type MC4100 membranes were injected with proOmpA or LamB signal peptides into Xenopus oocytes. However, such inhibition was lost when observed with oocyte-endogenous signal peptides in the absence of bacterial signal peptides. Moreover, azide did not show complete inhibition upon using SecYEG¯ membranes or SecYEG¯ reconstituted membranes plus excess SecA in the presence or absence of LamB signal peptides. Such conformational alterations reflect different sensitivity in response to azide during the opening of protein-conducting channels.

英文介系詞在句子裡所扮演的角色通常是用來使介系詞片語更精確地補述上下文，英文的母語使用者可以很直覺地使用。然而電腦不瞭解語義，因此不容易判斷介系詞修飾對象；非英文母語使用者則不容易直覺地使用正確的介系詞。所以本研究將專注於介系詞片語定位與介系詞推薦的議題。 在本研究將這二個介系詞議題抽象化為一個決策問題，並提出一個一般化的解決方法。這二個問題共通的部分在於動詞片語，一個簡單的動詞片語含有最重要的四個中心詞（headword）：動詞、名詞一、介系詞和名詞二。由這四個中心詞做為出發點，透過WordNet做階層式的選擇，在大量的案例中尋找語義上共通的部分，再利用機器學習的方法建構一般化的模型。此外，針對介系詞片語定的問題，我們挑選較具挑戰性介系詞做實驗。 藉由使用真實生活語料，我們的方法處理介系詞片語定位的問題，比同樣考慮四個中心詞的最大熵值法（Max Entropy）好；但與考慮上下文的Stanford剖析器差不多。而在介系詞推薦的問題裡，較難有全面比較的對象，但我們的方法精準度可達到53.14%。 本研究發現，高層次的語義可以使分類器有不錯的分類效果，而透過階層式的選擇語義能使分類效果更佳。這顯示我們確實可以透過語義歸納一套準則，用於這二個介系詞的議題。相信成果在未來會對機器翻譯與文本校對的相關研究有所價值。 / This thesis focuses on problems of attachment of prepositional phrases (PPs) and problems of prepositional suggestions. Determining the correct PP attachment is not easy for computers. Using correct prepositions is not easy for learners of English as a second language. I transform the problems of PPs attachment and prepositional suggestion into an abstract model, and apply the same computational procedures to solve these two problems. The common model features four headwords, i.e., the verb, the first noun, the preposition, and the second noun in the prepositional phrases. My methods consider the semantic features of the headwords in WordNet to train classification models, and apply the learned models for tackling the attachment and suggestion problems. This exploration of PP attachment problems is special in that only those PPs that are almost equally possible to attach to the verb and the first noun were used in the study. The proposed models consider only four headwords to achieve satisfactory performances. In experiments for PP attachment, my methods outperformed a Maximum Entropy classifier which also considered four headwords. The performances of my methods and of the Stanford parsers were similar, while the Stanford parsers had access to the complete sentences to judge the attachments. In experiments for prepositional suggestions, my methods found the correct prepositions 53.14% of the time, which is not as good as the best performing system today. This study reconfirms that semantic information is instrument for both PP attachment and prepositional suggestions. High level semantic information helped to offer good performances, and hierarchical semantic synsets helped to improve the observed results. I believe that the reported results are valuable for future studies of PP attachment and prepositional suggestions, which are key components for machine translation and text proofreading.

Este trabalho estuda o campo da edição de texto na produção editorial de livros no Brasil. O objetivo é apontar algumas distinções necessárias às pesquisas acadêmicas sobre edição de textos, e importantes para a formação de pesquisadores e profissionais. A proposta centra-se nas principais etapas (edição de texto, preparação de originais e revisão de provas) que compõem o processo por que passa um texto a ser transformado em livro. Parte-se de análise dos manuais de editoração e de pesquisas acadêmicas que tangenciam o tema. E articula-se um diálogo entre alguns trabalhos acadêmicos das áreas de psicolinguística e psicologia cognitiva sobre leitura em suas relações possíveis com os estudos em editoração e a experiência empírica dos editores de texto. Contribui-se, assim, para o aprimoramento dos conhecimentos sobre a edição de texto, entre eles: as etapas, os objetivos principais e secundários, as estratégias, as tarefas e um conjunto de elementos necessários para que um profissional atue como editor de texto consciente de sua intervenção. / It is the purpose of this thesis to study the field of text-editing in the Brazilian book publishing industry. Its main aim is to point out a few distinctions deemed necessary not only to the academic research but also to the formation of researchers and professionals in this particular field of study and work. In order to achieve this goal, the main phases the text goes through on its way to becoming a book -- text-editing, copy-editing and proofreading -- are here taken into account and characterized. Examining the available manuals and academic papers on the book editing field is the starting point of this research. It then goes on to promoting a dialogue between the book editing field and other areas of expertise, such as psycholinguistics and cognitive psychology, that also deal with the process of reading. The possible relationships this dialogue reveals and the actual praxis of text editing are then analyzed. Contributing to a better knowledge of what text-editing actually involves is therefore a primary aim of this thesis. It intends to shed some light on the specific phases, primary and secondary goals and strategies text-editing comprises, as well as on a series of elements needed not only for the praxis of text-editing itself but also for a more conscientious practice of this activity.

Orientador: Maria Viviane do Amaral Veras / Dissertação (mestrado) - Universidade Estadual de Campinas, Instituto de Estudos da Linguagem / Made available in DSpace on 2018-08-25T09:37:27Z (GMT). No. of bitstreams: 1 Pereira_JulianaCristinaFernandes_M.pdf: 1767935 bytes, checksum: cc5ac8a2f3da9afe0ab9dafb9186bb4b (MD5) Previous issue date: 2014 / Resumo: Nos últimos anos, têm sido divulgados inúmeros trabalhos com enfoque na representação do tradutor e do intérprete na literatura e no cinema, mas não tantos focam o revisor. E é justamente na fronteira entre realidade e ficção que se encontra o revisor-detetive Daniel Hernández, personagem central desta pesquisa, uma vez que seu autor Rodolfo Walsh (revisor, tradutor, ensaísta e, posteriormente, militante político) retira-o do mundo real e transporta-o para o ficcional. A presente pesquisa, portanto, tem por objetivo analisar a narrativa A aventura das provas de prelo, do escritor Rodolfo Walsh, como uma metáfora da tarefa do revisor, de seus rastros, de sua escrita, de seu trabalho de tradutor. Junto com o passo a passo do ofício do revisor, apresentamos o trajeto desta pesquisa: uma breve apresentação da vida de Rodolfo Walsh, bem como de sua literatura policial; conceitos relacionados à revisão, diretamente ligados ao dia a dia do revisor; a fundamentação teórica para a análise da tarefa do revisor e de sua criação como personagem de ficção, acompanhado de outros dois personagens: o revisor Raimundo Silva e o tradutor Gallus (abordados segundo uma visão mais conservadora, ou tradicional, de tradução e revisão) e o intérprete, senhor Kapasi (em cujas características reconhecemos uma visão denominada pós-estruturalista); a análise do personagem central criado por Rodolfo Walsh, o revisor-detetive Daniel Hernández. A título de desfecho, o ofício do revisor se apresenta como sempre mais ou menos terminado, sempre sujeito a falhas, confirmando que, ao menos no caso aqui estudado, não há crime perfeito / Abstract: Over the last years, a great number of papers have been written focusing on the part translators and interpreters play in literature and in the movies, but not many have concentrated on the proofreader's role. And precisely on the borderline between reality and fiction, we find proofreader-detective Daniel Hernández, the central character of this research, since his creator, author Rodolfo Walsh (proofreader, translator, essayist, and eventually, political activist), takes him out of the real world and plants him in the fictional. The scope of this paper, therefore, is to analyze the novel A aventura das provas de prelo (The Adventure of the Print Proofs), by author Rodolfo Walsh, as a metaphor of a proofreader¿s job, his tracks, his writings, and his work as a translator. Along with a step-by-step description of an proofreader¿s routine, we present the trajectory of this research: a brief presentation of the life of Rodolfo Walsh, as well as his police literature; concepts related to editing, directly connected to an proofreader¿s daily routine; the theoretical basis for the analysis of an proofreader¿s job and his creation as a fictional character, accompanied by two other characters: proofreader Raimundo Silva and translator Gallus (viewed through a more conservative or traditional lens of translation and editing) and the interpreter, Mr. Kapasi (in whose traits one can perceive a post-structuralist viewpoint); the analysis of the central character created by Rodolfo Walsh, proofreader-detective Daniel Hernández. By way of conclusion, the position of the proofreader always presents itself as more or less finished, always subject to flaws, confirming that, at least in the case studied here, there is no perfect crime / Mestrado / Mestra em Linguística Aplicada

Experiments have demonstrated that DNA molecules can compute like a machine to solve mathematical problems, which is significant because of their parallel computation ability. However, due to the nature of biochemical reactions, DNA computation suffers from errors, which are its main limitation. The abstract and kinetic Tile Assembly Models are now commonly used to simulate real DNA computing experiments, and to look for new methods to advance the accuracy of DNA-based computation. One means of controlling errors is through proofreading tile sets. Several such tile sets have been proposed in the literature, such as Chen and Goel’s snaked proofreading tile sets, the 2-way and 3-way overlay tile sets of Reif et al., and Rothemund and Cook’s n-way overlay tile sets. In the first part of this thesis, we analyze the performance of the Rothemund-Cook n-way overlay tile sets. We prove that the n-way overlay tile set contains n^2+3n+4 rule tiles. Simulation results show that these tile sets clearly perform better than tile sets without any error-control mechanism, and the performance improves as n increases. It is also proved that the error rates in assemblies formed by the 1-way and 2-way tile sets are O(epsilon^2), where epsilon is the error rate in assemblies without any error correction. In the second part of this thesis, we focus on a different error mechanism, namely,errors caused by imperfect or malformed tiles. We propose a model of malformed tiles, and consider the performance of various proofreading tile sets in the presence of malformed tiles. Our simulation results show that the Reif et al. 3-way overlay tile sets are able to best deal with malformed tiles. During the simulations, we observed that snaked proofreading tile sets always have trouble completing whole patterns when malformed tiles are present. We instead propose two modified snaked proofreading constructions, and verify through both simulations and analysis that the two modified constructions have much better performances. / Thesis (Master, Mathematics & Statistics) -- Queen's University, 2009-08-25 11:10:39.142