Call number: LD2668 .T4 1957 M68 / Master of Science

Branched-chain amino acid (BCAA) supplementation may improve recovery from competitive endurance training by reducing symptoms of central and peripheral fatigue. The purpose of this dissertation was to provide evidence for the use of BCAAs as a training nutrition strategy in order to improve recovery from training and further drive adaptive processes to training by increasing readiness to train. Collegiate distance runners undergoing intense competitive endurance training were monitored for symptoms of muscular soreness and psychological stress through a rated soreness chart and the Daily Analysis of Life Demands for Athletes Survey (DALDA) while taking either a BCAA supplement or a placebo. When on the BCAA supplement, athletes reported significantly fewer symptoms of psychological stress and reduced muscular soreness. These results point to the importance of nutrient bioavailability, specifically BCAAs, on recovery parameters when undergoing intensive training. This concept was also demonstrated in a case study on a trained distance runner who underwent intensive training for an ultra-endurance marathon. Running kinematics were assessed using Kinovea open-sourced software (Version 0.8.15) during a series of constant-paced endurance runs while on the BCAA supplement or a placebo. We observed a reduction in vertical oscillation when the runner was on the BCAA supplement, indicating improved muscle recovery and therefore efficiency of movement. Improving recovery by reducing global central and peripheral fatigue symptoms may increase readiness to train and further promote desired training adaptations.

Em 2005, Carolino e colaboradores isolaram uma fração neurotóxida de Averrhoa carambola (carambola) que foi capaz de apresentar sintomas em ratos e camundongos e foi chamada de AcTx. Acredita-se que o aminoácido denominado caramboxina esteja presente nesta fração e seja o metabólito secundário responsável por envenenamento pela ingestão da fruta em pacientes com insuficiência renal crônica. Apesar da estrutura da caramboxina ter sido elucidada por análises espectrométricas, a síntese deste composto visando caracterização estrutural e análise biológica ainda não havia sido realizada. Assim o objetivo deste trabalho foi investigar a síntese total da caramboxina visando sua caracterização estrutural e análise biológica dos compostos obtidos. Quatro rotas foram propostas, três das quais fizeram uso de reagentes organometálicos. Aquela que não fez uso de reagentes organometálicos foi a que se mostrou mais eficiente. / In 2005, Carolino and co-workers isolated a neurotoxic fraction of Averrhoa carambola (star fruit) that was able to cause seizures in rats and mice and was named AcTx. It is believed that the amino acid caramboxine, present in this fraction, is one of the secondary metabolites responsible for poisoning by fruit intake by patients with chronic renal failure. Despite its structure has been elucidated by spectrometric analysis, the synthesis of this compound aiming structural characterization and biological evaluation has not been still accomplished. Thus, the objective of this project was the total synthesis of caramboxina aiming their structural characterization and biological testing of the compounds obtained. Four routes have been proposed and investigated, three of them using organometallic reagents. The route that didn\'t use organometallic intermediates appeared to be the most efficient.

Thesis advisor: Abhishek Chatterjee / Förster Resonance Energy Transfer (FRET) offers a powerful approach to study biomolecular dynamics in vitro as well as in vivo. The ability to apply FRET imaging to proteins in living cells provides an excellent tool to monitor important dynamic events such as protein conformational changes, protein-protein interactions, and proteolysis reactions. However, selectively incorporating two distinct fluorophores into the target protein(s) that are capable of FRET interaction within the complex cellular milieu is challenging. Consequently, terminal fusion to genetically encoded fluorescent proteins has emerged as the predominant labeling strategy for FRET studies in vivo. However, a major limitation of this strategy stems from the large size of the fluorescent proteins, which may perturb the native properties of the target, and restricted attachment only to the termini of the target. We reasoned that using genetically encoded fluorescent unnatural amino acids would overcome several of these challenges associated with currently available labeling strategies owing to their small size and the ability to introduce them site- specifically and co-translationally. Here, we report the use of the fluorescent unnatural amino acid “Anap” as a FRET donor with green and yellow fluorescent protein acceptors. We demonstrate the utility of this labeling strategy using proteolysis and conformational change models, and step towards in vivo studies by further developing a proteolysis system in cell lysates. / Thesis (MS) — Boston College, 2016. / Submitted to: Boston College. Graduate School of Arts and Sciences. / Discipline: Chemistry.

<p>Phenylalanine (Phe) is a proteinogenic aromatic amino acid that also serves as a precursor for numerous primary and secondary metabolites in plants. Phe is synthesized from chorismate, the final product of the shikimate pathway. In plants, Phe is predominantly synthesized in the plastids via the arogenate pathway, while most Phe-derived compounds are produced in the cytoplasm, requiring exportation of Phe from plastids to the cytosol. Here, we provided genetic evidences that a<i> Petunia hybrida</i> plastidial cationic amino acid transporter (PhpCAT) participates in the exportation of Phe from plastids, as well as regulation of carbon flux through Phe biosynthesis.</p> <p> By using reverse genetics, we demonstrated that a petunia phenylpyruvate aminotransferase (PhPPY-AT) is able to convert phenylpyruvate to Phe in the cytosol <i>in vivo</i>, and that a cytosolic chorismate mutase (CM2), which converts chorismate to prephenate, directs carbon flux from the plastidial Phe biosynthesis pathway towards the cytosolic pathway. Downregulation of <i>PhPPY-AT</i> and <i>PhCM2</i> resulted in significant decreases in Phe levels and emission of Phe-derived volatiles in petunia flowers, respectively. Metabolic flux analysis showed that the carbon flux through the cytosolic Phe biosynthesis pathway is significantly lower in <i>PhCM2</i> RNAi petunia flowers relative to wild type control. We also demonstrated that the conversion of prephenate to phenylpyruvate in the cytosol is catalyzed by a cytosolic prephenate dehydratase (PDT) produced from an alternative transcription start site of a known plastidial arogenate dehydratase (ADT). These results suggest that a microbial-like phenylpyruvate pathway for Phe biosynthesis operates in the cytosol of plant cells and the cytosolic pathway splits from the plastidial pathway at chorismate.</p> <p> To evaluate the metabolic potential of the cytosolic phenylpyruvate pathway, <i>PhCM2 </i>overexpressing transgenic petunia plants were generated. Unexpectedly, Phe levels and emission of Phe-derived volatiles were both reduced, even though the flux through the cytosolic pathway was increased relative to wild type control. Electron microscopy, metabolic profiling and metabolic flux analysis revealed that the number of leucoplasts, starch levels and flux through the plastidial pathway were all reduced in <i>PhCM2</i> overexpression lines, while the concentrations of auxin and its biosynthetic intermediate, indole-3-pyruvic acid (IPA), were elevated. Overexpression of Arabidopsis aminotransferase VAS1, which converts IPA to Trp, in <i>PhCM2</i> overexpression petunia background recovered Phe levels and Phe-derived volatiles emission. These results indicate that there exists a metabolic crosstalk between cytosolic Phe production and Trp-dependent auxin biosynthesis .</p> <p> Our research completed the post-chorismate cytosolic Phe biosynthesis pathway in plants and revealed possible metabolic crosstalk between cytosolic Phe production and auxin biosynthesis in plant cells, providing targets for future genetic modification of metabolites in plants.</p>

Objetivo. Amaranto é considerado um alimento funcional devido às suas propriedades de redução de colesterol plasmático. Um possível componente do amaranto responsável por este efeito é a proteína.Métodos. Neste estudo, foi produzido isolado protéico de amaranto através da solubilização da proteína em pH 11 e precipitação em pH 5,7, obtendo-se o isolado com pureza de 96% de proteína. Este isolado protéico foi utilizado como fonte de proteínas em dietas experimentais para hamsters que tiveram hipercolesterolemia induzida, previamente, por dieta contendo 30% de caseína e 0,05% de colesterol, durante 3 semanas. Os animais foram, então, distribuídos em três grupos (n=11 animais/grupo) e foram alimentados com dietas contendo: (A) 20% caseína (controle), (B) 20% proteína de amaranto purificada (grupo substituição) e (C) 20% caseína + 10% proteína de amaranto purificada (grupo suplementação). Resultados. Comparando-se com a dieta controle, o grupo da suplementação e o da substituição tiveram dramáticas reduções do nível de colesterol plasmático, 30% (p<0,05) e 51% (p<0,05) respectivamente, enquanto o controle apresentou redução de apenas 7% após os 28 dias de dieta. Já na primeira semana este comportamento de redução para as duas dietas contendo amaranto foi percebido, e a redução foi mais marcante na fração LDL. Os mecanismos envolvidos na redução do colesterol plasmático foram investigados. A digestibilidade verdadeira da proteína do amaranto foi igual à da caseína. A excreção de ácidos biliares foi inversamente proporcional à redução do colesterol plasmático nas diferentes dietas, enquanto que o colesterol excretado foi proporcional à redução do colesterol. Quando aminoácidos livres simulando o perfil da proteína de amaranto foram utilizados como única fonte de nitrogênio da dieta, a redução dos níveis de colesterol foi de 11%. A dieta contendo caseína e suplementada com arginina de forma a resultar numa relação lisina/arginina de 0,5 (a mesma observada na proteína de amaranto), mostrou-se deletéria aos parâmetros plasmáticos. Conclusões. Comprovou-se que a proteína de amaranto reduz o colesterol plasmático. A digestibilidade e excreção de ácidos biliares não estão relacionados com a redução do colesterol provocada pela proteína do amaranto. A relação dos aminoácidos lisina/arginina explica apenas parcialmente o mecanismo e apenas a proteína íntegra tem efeito sobre a excreção de colesterol nas fezes. O mecanismo envolvido na redução do colesterol nestes experimentos ainda não está totalmente elucidado, sugerindo a necessidade de futuros estudos da ação direta de peptídeos formados pela digestão incompleta da proteína do amaranto no metabolismo lipídico. / Objective. Amaranth has been considered a functional food because its consumption can lower blood cholesterol levels. In the present work the effect of amaranth protein on this property was investigated in hamsters. A possible component in amaranth grain that would respond for this effect is the protein fraction. Methods. In this study the amaranth protein was isolated by its alkaline solubilization at pH 11 and acid precipitation at pH 5.7. The isolate thus produced was defatted and resulted in a protein content of about 96%. This product was introduced in experimental diets to fed hamsters that previously had their blood cholesterol increased by a diet containing 30% casein and 0.05% cholesterol during 3 weeks Animals were then, divided in 3 groups (n = 11/group) were fed diets containing (g/100 g diet): (A) 20 casein (control), (B) 20 purified amaranth’s protein (group replacement), (C) 20 casein + 10 purified amaranth’s protein (group supplementation) for 4 wks. Results. The results showed that when amaranth was the sole protein source (at 20% level) or it was admixed with casein (20% casein +10% of amaranth protein), the hypercholesterolemized hamsters had a significant (P < 0.05) reduction in cholesterol levels (51 and 30%, respectively) as compared 7% reduction of the control group (20% casein). In the first week of diet the decrease was already observed. The lowering was mainly in LDL fraction. The mecanisms involved in lowering plasma cholesterol were investigated. Digestibility of amaranth protein was as high. The bile acids excretion was inversely proportional to plasma cholesterol lowering, while cholesterol excretion in feces was directly proportional. When free amino acids simulating the amaranth protein were used as the only nitrogen source of diet the cholesterol reduction was about 11%. Casein supplemented with arginine to bring the lysine/arginine ratio to 0.5, as observed in amaranth protein, was had deleterious effects to hamsters’ cholesterol levels. The bile acid and cholesterol excretion of this trial were equal to all groups. Conclusions. Amaranth´s protein reduces plasma cholesterol. Digestibility and bile acid excretion are not related to hypocholesterolemic effect of amaranth’s protein. The proportion between lysine/argine is a partial explanation for this effect, but the presence of whole protein is necessary for the higher cholesterol excretion in feces. The full understanding of mechanisms involved in cholesterol reduction in these experiments is not fully elucidated, suggesting further research on the direct action in lipid metabolism by peptides originated from the incomplete digestion of amaranth protein.

by Ho, Wai Fun. / Thesis (M.Sc.)--Chinese University of Hong Kong, 1994. / Includes bibliographical references (leaves 79-87). / LIST OF TABLES --- p.iv / LIST OF FIGURES --- p.vi / ACKNOWLEDGEMENTS --- p.vii / ABSTRACT --- p.viii / Chapter 1. --- INTRODUCTION --- p.1 / Chapter 1.1 --- METABOLIC DERANGEMENTS AND CACHEXIA IN CANCER --- p.1 / Chapter 1.2 --- PROTEIN METABOLISM IN MALIGNANCY --- p.4 / Chapter 1.3 --- REVIEW OF REPORTS ON AMINO ACID DISTURBANCES IN MALIGNANCY --- p.5 / Chapter 1.4 --- AMINO ACID ANALYSIS BY HIGH PERFORMANCE LIQUID CHROMATOGRAPHY --- p.10 / Chapter 1.4.1 --- Amino Acid Analysis by Ion-Exchange HPLC --- p.11 / Chapter 1.4.2 --- Amino Acid Analysis by Reversed-Phase HPLC --- p.13 / Chapter 1.4.3 --- Derivatizing Agents --- p.15 / Chapter 1.5 --- CHOICE OF CANCER PATIENTS AND METHODOLOGY FOR THIS STUDY --- p.19 / Chapter 1.5.1 --- Choice of Cancer Patients --- p.19 / Chapter 1.5.2 --- Methodology Chosen and Its Principle --- p.20 / Chapter 2. --- OBJECTIVES --- p.23 / Chapter 3. --- MATERIALS AND METHODS --- p.24 / Chapter 3.1 --- STUDY SUBJECTS --- p.24 / Chapter 3.1.1 --- Patients --- p.24 / Chapter 3.1.2 --- Control Subjects --- p.25 / Chapter 3.2 --- CLINICAL FEATURES --- p.25 / Chapter 3.3 --- BLOOD COLLECTION --- p.25 / Chapter 3.4 --- GENERAL BIOCHEMICAL TESTS --- p.26 / Chapter 3.5 --- PLASMA AMINO ACID ANALYSIS BY HPLC --- p.26 / Chapter 3.5.1 --- Apparatus --- p.26 / Chapter 3.5.2 --- Reagents --- p.27 / Chapter 3.5.3 --- Reagent Preparation --- p.28 / Chapter 3.5.3.1 --- Mobile phase --- p.28 / Chapter 3.5.3.2 --- Derivatizing reagent --- p.29 / Chapter 3.5.4 --- Standard Preparation --- p.29 / Chapter 3.5.4.1 --- Internal standard solution --- p.29 / Chapter 3.5.4.2 --- Composite standard solution --- p.30 / Chapter 3.5.4.3 --- Composite standard-internal standard mixture --- p.32 / Chapter 3.5.5 --- Sample Preparation --- p.32 / Chapter 3.5.5.1 --- Protein removal --- p.32 / Chapter 3.5.5.2 --- Addition of internal standard --- p.32 / Chapter 3.5.6 --- Preparation of Samples for the WISP Sample Processor --- p.33 / Chapter 3.5.7 --- Sample Queue for the WISP Sample Processor --- p.33 / Chapter 3.5.8 --- Automated Derivatization Procedure --- p.36 / Chapter 3.5.9 --- Chromatographic Conditions --- p.36 / Chapter 3.6 --- STATISTICAL STUDIES --- p.38 / Chapter 4. --- RESULTS --- p.40 / Chapter 4.1 --- ANALYTICAL PERFORMANCES --- p.40 / Chapter 4.1.1 --- Chromatograms --- p.40 / Chapter 4.1.2 --- Precision --- p.47 / Chapter 4.1.3 --- Linearity --- p.47 / Chapter 4.1.4 --- Analytical Recovery --- p.51 / Chapter 4.2 --- DATA DISTRIBUTION STUDIES --- p.53 / Chapter 4.3 --- PATIENTS' ANTHROPOMETRIC DATA AND BLOOD BIOCHEMISTRY --- p.55 / Chapter 4.4 --- FREE PLASMA AMINO ACID CONCENTRATIONS IN NORMAL CONTROLS --- p.59 / Chapter 4.5 --- FREE PLASMA AMINO ACID CONCENTRATIONS IN CANCER PATIENTS --- p.59 / Chapter 5. --- DISCUSSION --- p.68 / Chapter 5.1 --- METHOD ESTABLISHMENT --- p.68 / Chapter 5.2 --- NORMAL CONTROLS --- p.68 / Chapter 5.3 --- CANCER PATIENTS --- p.71 / Chapter 5.3.1 --- Nasopharyngeal Cancer --- p.71 / Chapter 5.3.2 --- Lung Cancer --- p.71 / Chapter 5.3.3 --- Breast Cancer --- p.72 / Chapter 5.3.4 --- Colorectal Cancer --- p.74 / Chapter 5.4 --- SUMMARY OF THE PLASMA AMINO ACID PROFILES IN CANCER --- p.75 / Chapter 5.5 --- FURTHER STUDIES --- p.76 / Chapter 6. --- CONCLUSION --- p.78 / Chapter 7. --- REFERENCES --- p.79

Poon Wai Mei Emily. / Thesis (M.Phil.)--Chinese University of Hong Kong, 1999. / Includes bibliographical references (leaves 74-81). / Abstracts in English and Chinese. / Chapter Chapter 1: --- Introduction --- p.1 / Chapter 1.1. --- Importance of amino acids --- p.1 / Chapter 1.1.1. --- Clinical samples --- p.1 / Chapter 1.1.2. --- Food samples --- p.2 / Chapter 1.2. --- Reviews of amino acid analysis --- p.6 / Chapter 1.2.1. --- Ion-exchange chromatography --- p.6 / Chapter 1.2.2. --- Gas chromatography --- p.6 / Chapter 1.2.3. --- Thin layer chromatography --- p.8 / Chapter 1.2.4. --- Flow injection analysis --- p.8 / Chapter 1.2.5. --- Liquid chromatography --- p.9 / Chapter 1.2.6. --- Capillary electrophoresis --- p.10 / Chapter 1.2.7. --- Methods of detecting amino acid without derivatization --- p.11 / Chapter 1.3. --- Determination of amino acids by reversed-phase ion-pair chromatography --- p.12 / Chapter 1.4. --- The objectives of the study --- p.15 / Chapter Chapter 2: --- Experimental --- p.16 / Chapter 2.1. --- Materials --- p.16 / Chapter 2.2. --- Apparatus --- p.16 / Chapter 2.3. --- Samples --- p.16 / Chapter 2.4. --- Procedures --- p.17 / Chapter 2.4.1. --- Preparation of amino acid standard solution (stock solutions) --- p.17 / Chapter 2.4.2. --- Method development --- p.17 / Chapter 2.4.3. --- Samples preparation --- p.18 / Chapter 2.4.4. --- Preparation of Dowex column --- p.18 / Chapter 2.4.5. --- Extraction of amino acids from samples --- p.19 / Chapter 2.4.6. --- Recovery test --- p.20 / Chapter Chapter 3: --- Results and Discussions --- p.21 / Chapter 3.1. --- Optimization --- p.21 / Chapter 3.1.1. --- pH --- p.21 / Chapter 3.1.2. --- Ion-interacting reagent --- p.22 / Chapter 3.1.3. --- Organic solvent --- p.29 / Chapter 3.1.4. --- Temperature --- p.34 / Chapter 3.1.5. --- Chromatographic conditions --- p.36 / Chapter 3.2. --- Application --- p.45 / Chapter 3.2.1. --- Precision of injection --- p.45 / Chapter 3.2.2. --- Accuracy of the method --- p.46 / Chapter 3.2.3. --- The concentration of amino acids in food samples --- p.50 / Chapter 3.2.3.1. --- Citrus fruits --- p.50 / Chapter 3.2.3.2. --- Orange juice drinks --- p.60 / Chapter 3.2.3.3. --- Chinese honey --- p.65 / Chapter 3.2.3.4. --- New Zealand honey --- p.67 / Chapter 3.2.3.5. --- Energy drinks --- p.70 / Chapter Chapter 4 : --- Conclusion --- p.72 / Chapter Chapter 5 : --- Bibliographies --- p.74 / Chapter 6. --- Appendices --- p.82 / Chapter 6.1. --- Table 1 : Ingredients of orange juice drinks --- p.82 / Chapter 6.2. --- Table 2 : Honey samples --- p.83 / Chapter 6.3. --- Table 3 : Ingredients of energy drinks --- p.83

No description available.

The ribosome is responsible for mRNA-templated protein translation in all living cells. The translational machinery (TM) has evolved to use 20 amino acids each esterified onto one of several tRNA bodies. While the active site of the ribosome, known as the peptidyl transferase center (PTC), is able to handle a remarkable amount of substrate diversity, many classes of unnatural amino acids are not compatible with the TM. For example, in the field of unnatural amino acid mutagenesis, the site-specific incorporation of biologically useful amino acids into proteins, such as fluorophores, has often proven to be unfeasible. This runs counter to the accepted notion that the ribosome is blind to the structure of the amino acid and is capable of accepting any amino acid as long as the mRNA codon: tRNA anticodon pairing is correct. Two studies by our group set out to test the hypothesis that the ribosome can indeed discriminate the structure of the amino acid. Using a fully purified E. coli translation system, the first study showed that natural amino acids misacylated onto fully modified but non-native tRNAs show small but reproducible effects on the steps of aminoacyl-tRNA (aa-tRNA) selection. The second study, in which I participated, utilized D-aa-tRNAs in the same E. coli translation system to study how amino acids of the inverted stereochemistry to those found in ribosomally-synthesized proteins affect translation elongation. We showed that these unnatural substrates serve as peptidyl acceptors but once translocated into the P-site of the ribosome, fail as peptidyl donors and stall translation elongation by inactivating the PTC. The motivation of my work has been to further characterize the effects of D-aa-tRNAs on translation elongation. To this end, I examined how the PTC is affected structurally and functionally by the presence of ribosomal substrates containing D-amino acids. Chapter one contains an introduction to this work. Chapter two describes chemical probing experiments that demonstrate that the presence of peptidyl-D-aminoacyl-tRNAs in the P-site of the ribosome allosterically modulates the secondary structure of ribosomal exit tunnel nucleotides A2058 and A2059. Chapter three describes how the reactivity of peptidyl-D-aminoacyl-tRNAs to form tripeptides is highly dependent on the identity of the amino acid it is reacting with; protein yields can be close to what is obtained with natural amino acids or almost completely abolished. Chapter four contains the methods used to do this research. From the observations presented here as well as from the work of other laboratories, a picture of the PTC emerges in which the pairing of the A- and P- site substrates is integral in either promoting or suppressing catalysis by the PTC. This work has implications for the field of unnatural amino acid mutagenesis, particularly for strategies to improve the incorporation of interesting unnatural amino acid by the ribosome. In addition, this work adds an important aspect to the growing body of knowledge of ribosome stalling at the PTC.