A ribosomal system has been derived from <u>Cellvibrio gilus</u> which is active with respect to amino acid incorporation into hot trichloroacetic acid-insoluble material. Optimal incorporation systems showed a requirement for ribosomes, pH 5 enzyme protein, ATP plus ATP-generating system, magnesium ion, ammonium ion, and GTP. ¹⁴C-phenylalanine incorporation was stimulated by poly U in the presence of t-RNA which has been derived from this bacterium. The t-RNA had some activation effect on poly U-directed ¹⁴C-phenylalanine incorporation, whereas no effect on natural m-RNA primed ¹⁴C-leucine incorporation was observed. A complementary unlabeled amino acid mixture inhibited ¹⁴C-leucine incorporation. ¹⁴C-leucine incorporation was greatly inhibited by RBAase and puromycin, moderately inhibited by chloramphenicol, but not effected significantly by DNAase and actinomycin D. The optimum pH and temperature for ¹⁴C-leucine incorporation were 7.8 and 300° C, respectively. Polyamiaes (putrescine, spermine, and spermidine) had a slight stimulatory effect on poly U-directed.¹⁴C-phenylalanine incorporation, but no effect on ¹⁴C-Leucine incorporation. / Master of Science

No / Despite the extensive utilization of biomolecule-titania interfaces, biomolecular recognition and interactions at the aqueous titania interface remain far from being fully understood. Here, atomistic molecular dynamics simulations, in partnership with metadynamics, are used to calculate the free energy of adsorption of different amino acid side chain analogues at the negatively-charged aqueous rutile TiO2 (110) interface, under conditions corresponding with neutral pH. Our calculations predict that charged amino acid analogues have a relatively high affinity to the titania surface, with the arginine analogue predicted to be the strongest binder. Interactions between uncharged amino acid analogues and titania are found to be repulsive or weak at best. All of the residues that bound to the negatively-charged interface show a relatively stronger adsorption compared with the charge-neutral interface, including the negatively-charged analogue. Of the analogues that are found to bind to the titania surface, the rank ordering of the binding affinities is predicted to be "arginine" > "lysine" ≈ aspartic acid > "serine". This is the same ordering as was found previously for the charge-neutral aqueous titania interface. Our results show very good agreement with available experimental data and can provide a baseline for the interpretation of peptide-TiO2 adsorption data. / Veski

Master of Science / Department of Animal Sciences and Industry / Michael D. Tokach / A total of 5,212 nursery pigs were used in 11 experiments to evaluate amino acids in nursery pig diets. Experiments 1 and 2 were conducted to determine whether the Lys level fed during one phase of the nursery influenced the response to Lys during subsequent phases. Experiment 1 tested a wide range of dietary Lys in 2 phases and reported that pigs fed high Lys during each period had increased growth performance; however, compensatory growth occurred for the pigs previously fed low Lys diets, resulting in no impact on overall ADG or final BW. Experiment 2 tested a narrow range of dietary Lys in 3 phases and found that marginally deficient diets can be fed in the early nursery phases without influencing final BW or the response to Lys levels in subsequent phases. Both experiments demonstrate that the low dietary Lys levels used in each can be fed in the early nursery phases with no negative impact on overall nursery growth rate provided that adequate levels are fed thereafter. Experiments 3 to 6 were conducted to determine the standardized ileal digestible (SID) Lys requirement of nursery pigs from 7- to 14-kg. Data from all experiments were combined and break-point and quadratic broken-line analysis was used to determine the estimated SID Lys requirement. The SID Lys requirement for optimal growth was at least 1.30% for ADG and 1.37% for G:F, or at least 3.86 and 4.19 g SID Lys/Mcal ME, respectively. Experiments 7 to 11 were conducted to evaluate the effect of replacing specialty protein sources with crystalline AA and AA requirements for 7- to 12-kg pigs. Experiment 7 demonstrated that crystalline AA can be used to replace fish meal in diets with no negative effects on growth performance. Experiment 8 demonstrated that L-Trp, L-Val, and a source of non-essential AA were needed in low-CP, AA-fortified nursery diets to achieve maximum growth performance, whereas the addition of L-Ile was not required. Experiment 9 indicated that feeding greater than 7.35% total Lys:CP decreased growth performance and Exp. 10 indicated that a SID Val:Lys ratio of 65% was sufficient for optimal growth of early nursery pigs. Implementing the results from the previous experiments, Exp. 11 determined that crystalline AA in nursery pigs diets can replace high amounts of fish meal, meat and bone meal, and poultry meal when balanced for minimum AA ratios and maximum Lys:CP with no negative effect on growth performance.

Studies were evaluated on the effects of known growth factors on the growth and carotenogenesis of Corynebacterium species strain 7ElC. The complex medium, Tryptic Soy Broth,was found to stimulate growth and production of more pigment in the light and in the dark than did a mineral salts-glucose medium. A complete amino acid mixture added to LSG enhanced carotenogenesis in the dark in Corynebacterium 7ElC, while B-vitamins retarded carotenogenesis. No absolute requirement for one or more amino acids was found,indicating a multiple amino acid requirement. The fewest amino acids found to stimulate carotenogenesis in the dark were a combination of those in the Serine and Histidine families which include serine, glycine, cysteine, and histidine.

by Lui Ka Man. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2003. / Includes bibliographical references (leaves 143-166). / Abstracts in English and Chinese. / Acknowledgements --- p.i / List of Abbreviations --- p.ii / Abstract --- p.iv / 摘要 --- p.viii / List of Tables --- p.xi / List of Figures --- p.xii / Contents --- p.xvii / Chapter CHAPTER 1 --- INTRODUCTION / Chapter 1.1 --- Sulfur-Containing Amino Acids --- p.1 / Chapter 1.2 --- Metabolism of Sulfur-Containing Amino Acids --- p.3 / Chapter 1.2.1 --- The Metabolic Pathways --- p.3 / Chapter 1.2.2 --- Metabolic Regulation of Homocysteine --- p.7 / Chapter 1.3 --- Sulfur-Containing Amino acids and Health --- p.10 / Chapter 1.3.1 --- Sulfur-Containing Amino Acids and Renal Diseases --- p.11 / Chapter 1.3.2 --- Sulfur-Containing Amino Acids and the Nervous System --- p.13 / Chapter 1.3.3 --- Sulfur-Containing Amino Acids and Heart Diseases --- p.16 / Chapter 1.3.4 --- Sulfur-Containing Amino Acids and Liver Diseases --- p.20 / Chapter 1.4 --- Vitamin C (Ascorbic Acid) --- p.21 / Chapter 1.4.1 --- Biosynthesis 、 --- p.23 / Chapter 1.4.2 --- Vitamin C Transport System --- p.24 / Chapter 1.4.2.1 --- Uptake of Ascorbic Acid --- p.25 / Chapter 1.4.2.2 --- Uptake of Dehydroascorbic Acid --- p.26 / Chapter 1.5 --- Vitamin C and Health --- p.28 / Chapter 1.5.1 --- Vitamin C and Heart Diseases --- p.28 / Chapter 1.5.2 --- Vitamin C and Renal Diseases --- p.29 / Chapter 1.5.3 --- Vitamin C and Brain Diseases --- p.30 / Chapter 1.5.4 --- Vitamin C and Liver Diseases --- p.31 / Chapter 1.6 --- The Aims of Project --- p.33 / Chapter CHAPTER 2 --- MATERIALS AND METHODS / Chapter 2.1 --- Materials --- p.35 / Chapter 2.2 --- Preparation of Materials --- p.37 / Chapter 2.2.1 --- Mobile Phase for Ascorbic Acid Analysis --- p.37 / Chapter 2.2.2 --- Mobile Phase for Amino Acid Analysis --- p.37 / Chapter 2.2.3 --- Reagents for RNA Extraction and Reverse Transcription- Polymerase Chain Reaction (RT-PCR) --- p.38 / Chapter 2.2.4 --- Sense and Antisense Primers --- p.38 / Chapter 2.2.5 --- Reagents for Electrophoresis --- p.40 / Chapter 2.3 --- Animals --- p.40 / Chapter 2.4 --- Experimental Studies --- p.41 / Chapter 2.5 --- Methods --- p.41 / Chapter 2.5.1 --- Ascorbic Acid Analysis --- p.41 / Chapter 2.5.1.1 --- Extraction of Ascorbic Acid --- p.41 / Chapter 2.5.1.2 --- Chromatography --- p.42 / Chapter 2.5.2 --- Amino Acid Analysis --- p.45 / Chapter 2.5.2.1 --- Sample Preparation --- p.45 / Chapter 2.5.2.2 --- Chromatography --- p.45 / Chapter 2.5.3 --- Determination of RNA Expression by RT-PCR Analysis --- p.50 / Chapter 2.5.3.1 --- RNA Isolation --- p.50 / Chapter 2.5.3.2 --- Measurement of RNA Yield --- p.51 / Chapter 2.5.3.3 --- Reverse Transcription --- p.51 / Chapter 2.5.3.4 --- Polymerase Chain Reaction (PCR) --- p.52 / Chapter 2.5.3.5 --- Separation of PCR Products by Agarose Gel Electrophoresis --- p.52 / Chapter 2.5.3.6 --- Quantification of Band Density --- p.53 / Chapter 2.6 --- Statistical Analysis --- p.53 / Chapter CHAPTER 3 --- RESULTS / Chapter 3.1 --- Effects of Sulfur-Containing Amino Acids and Leucine on Ascorbic Acid Distributions in Mice --- p.54 / Chapter 3.1.1 --- Effects of Sulfur-Containing Amino Acids and Leucine on Ascorbic Acid Concentrations in the Plasma --- p.55 / Chapter 3.1.2 --- Effects of Sulfur-Containing Amino Acids and Leucine on Ascorbic Acid Concentrations in the Kidney --- p.57 / Chapter 3.1.3 --- Effects of Sulfur-Containing Amino Acids and Leucine on Ascorbic Acid Concentrations in the Liver --- p.59 / Chapter 3.1.4 --- Effects of Sulfur-Containing Amino Acids and Leucine on Ascorbic Acid Concentrations in the Brain --- p.61 / Chapter 3.1.5 --- Effects of Sulfur-Containing Amino Acids and Leucine on Ascorbic Acid Concentrations in the Heart --- p.63 / Chapter 3.1.6 --- Summary --- p.65 / Chapter 3.2 --- Time-Response --- p.66 / Chapter 3.2.1 --- Effects of Sulfur-Containing Amino Acids and Leucine Administration for Various Periods on Ascorbic Acid Concentrations in the Plasma --- p.67 / Chapter 3.2.2 --- Effects of Sulfur-Containing Amino Acids and Leucine Administration for Various Periods on Ascorbic Acid Concentrations in the Kidney --- p.69 / Chapter 3.2.3 --- Effects of Sulfur-Containing Amino Acids and Leucine Administration for Various Periods on Ascorbic Acid Concentrations in the Liver --- p.71 / Chapter 3.2.4 --- Effects of Sulfur-Containing Amino Acids and Leucine Administration for Various Periods on Ascorbic Acid Concentrations in the Brain --- p.73 / Chapter 3.2.5 --- Effects of Sulfur-Containing Amino Acids and Leucine Administration for Various Periods on Ascorbic Acid / Chapter 3.2.6 --- Summary --- p.77 / Chapter 3.3 --- Effects of Sulfur-Containing Amino Acids and Leucine on SVCT and GLUT3 Gene Expressions --- p.78 / Chapter 3.3.1 --- Effects of Sulfur-Containing Amino Acids and Leucine Administration on SVCT mRNA Expression in the Kidney --- p.79 / Chapter 3.3.2 --- Effects of Sulfur-Containing Amino Acids and Leucine Administration on SVCT mRNA Expression in the Liver --- p.89 / Chapter 3.3.3 --- Effects of Sulfur-Containing Amino Acids and Leucine Administration on SVCT and GLUT3 mRNA Expression in the Brain --- p.98 / Chapter 3.3.4 --- Effects of Sulfur-Containing Amino Acids and Leucine Administration on SVCT mRNA Expression in the Heart --- p.109 / Chapter 3.3.5 --- Summary --- p.115 / Chapter 3.4 --- Sulfur-Containing Amino Acids Concentrations in the Plasma --- p.117 / Chapter 3.4.1 --- Effects of Administration of Sulfur-Containing Amino Acids and Leucine on Methionine Concentrations in the Plasma --- p.117 / Chapter 3.4.2 --- Effects of Administration of Sulfur-Containing Amino Acids and Leucine on Cystine Concentrations in the Plasma --- p.119 / Chapter 3.4.3 --- Effects of Administration of Sulfur-Containing Amino Acids and Leucine on Taurine Concentrations in the Plasma --- p.119 / Chapter 3.4.4 --- Effects of Administration of Sulfur-Containing Amino Acids and Leucine on Leucine Concentrations in the Plasma --- p.122 / Chapter 3.4.5 --- Summary --- p.124 / Chapter CHAPTER 4 --- DISCUSSION AND CONCLUSIONS / Chapter 4.1 --- Effects of Sulfur-Containing Amino Acids and Leucine on Ascorbic Acid Concentrations in Mice --- p.126 / Chapter 4.2 --- Effects of Sulfur-Containing Amino Acids and Leucine on SVCT and GLUT3 Gene Expressions --- p.131 / Chapter 4.3 --- Sulfur-Containing Amino Acids Concentrations in the Plasma --- p.136 / Chapter 4.4 --- Conclusions --- p.140 / REFERENCES --- p.143

Wong, Kin Sing. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2009. / Includes bibliographical references (leaves 86-98). / Abstract also in Chinese. / THESIS COMMITTEE --- p.i / ABSTRACT (ENGLISH) --- p.ii / ABSTRACT (CHINESE) --- p.iv / ACKNOWLEDGEMENTS --- p.v / DECLARATION --- p.vi / ABBREVIATIONS --- p.vii / TABLE OF CONTENTS --- p.x / LIST OF TABLES --- p.xiv / LIST OF FIGURES --- p.xv / Chapter CHAPTER 1 --- INTRODUCTION / Chapter 1.1. --- Antibiotics market and β-lactam antibiotics --- p.1 / Chapter 1.2. --- Semi-synthetic cephems --- p.1 / Chapter 1.3. --- Conversion of CPC to 7-ACA --- p.3 / Chapter 1.4. --- Chemical production versus enzymatic bioconversion --- p.5 / Chapter 1.5. --- Industrial two-step bioconversion of CPC --- p.11 / Chapter 1.6. --- Phylogenetics and physiological roles of DAAO --- p.15 / Chapter 1.7. --- Yeast DAAOs are suitable candidates for enzymatic bioconversion --- p.17 / Chapter 1.8. --- Structural and mechanistic studies of DAAOs --- p.18 / Chapter 1.9. --- "Modifications of pkDAAO, RgDAAO and TvDAAO" --- p.25 / Chapter 1.10. --- Objectives of the study --- p.26 / Chapter CHAPTER 2 --- HOMOLOGY MODELLING / Chapter 2.1. --- Introduction --- p.27 / Chapter 2.2. --- Methods / Chapter 2.2.1. --- Sequence alignment and selection of homologs --- p.28 / Chapter 2.2.2. --- Generation of three-dimensional TvDAAO model --- p.28 / Chapter 2.3. --- Results --- p.29 / Chapter 2.4. --- Discussion --- p.33 / Chapter CHAPTER 3 --- "MUTAGENESIS, EXPRESSION, PURIFICATION AND SCREENING OF MUTANTS" / Chapter 3.1. --- Introduction --- p.38 / Chapter 3.2. --- Materials and methods / Chapter 3.2.1. --- Cloning of TvDAAO mutants / Chapter 3.2.1.1. --- Preparation of competent E. coli --- p.39 / Chapter 3.2.1.2. --- Transformation of E. coli --- p.40 / Chapter 3.2.1.3. --- Agarose gel electrophoresis and gel-purification --- p.41 / Chapter 3.2.1.4. --- Plasmid extraction --- p.42 / Chapter 3.2.1.5. --- Site-directed mutagenesis of TvDAAO --- p.42 / Chapter 3.2.2. --- Heterologous expression and purification of mutants / Chapter 3.2.2.1 --- Shake flask fermentation --- p.45 / Chapter 3.2.2.2. --- Cell harvest and disruption --- p.45 / Chapter 3.2.2.3. --- Purification of WT and mutants --- p.47 / Chapter 3.2.2.4. --- Determination of protein concentration --- p.47 / Chapter 3.2.2.5. --- SDS-PAGE --- p.48 / Chapter 3.2.3. --- Screening of mutants --- p.48 / Chapter 3.3. --- Results / Chapter 3.3.1. --- Preparation of purified TvDAAO mutants --- p.50 / Chapter 3.3.2. --- Evaluation of activity and thermostability --- p.50 / Chapter 3.4. --- Discussion --- p.53 / Chapter CHAPTER 4 --- ENZYME KINETICS / Chapter 4.1. --- Introduction --- p.57 / Chapter 4.2. --- Materials and methods / Chapter 4.2.1. --- Standard assay --- p.58 / Chapter 4.2.2. --- Determination of kinetic parameters --- p.59 / Chapter 4.2.3. --- Inhibitory studies --- p.59 / Chapter 4.2.4. --- Effects of pH --- p.60 / Chapter 4.2.5. --- Heat treatments --- p.60 / Chapter 4.2.6. --- CD measurements --- p.61 / Chapter 4.3. --- Results / Chapter 4.3.1. --- Time progress curve analysis --- p.61 / Chapter 4.3.2. --- Kinetics of WT and mutants --- p.62 / Chapter 4.3.3. --- Temperature-dependent and time-dependent thermostability --- p.67 / Chapter 4.3.4. --- Secondary structure measurements --- p.71 / Chapter 4.4. --- Discussion --- p.71 / Chapter CHAPTER 5 --- CONCLUSIONS AND PERSEPECTIVES --- p.83 / BIBLIOGRAPHY --- p.86

Foi desenvolvido um conjunto de peças plásticas que permitem a montagem e representação dos aminoácidos mais comuns, bem como a construção de estruturas protéicas. Durante e após o desenvolvimento o material foi submetido a várias etapas de avaliação por professores (do ensino básico e universitário), alunos de pós-graduação e de graduação. A primeira etapa foi o desenvolvimento dos modelos em ambiente computacional, seguida da prototipagem das peças. Após discussão com a comunidade científica (apresentados na XXXVI Reunião Anual da SBBq em 2007) as sugestões foram implementadas nos modelos computacionais. Quatro moldes para injeção termoplástica foram projetados, detalhados e construídos, sob nossa orientação. As peças representando as estruturas que compõe os aminoácidos e ligações foram produzidas em grande escala e iniciou-se o processo final de avaliação. As peças apresentaram boas relações geométricas com as fórmulas estruturais dos aminoácidos obtidas de bancos de dados e livros didáticos. As conexões C&alpha;-amina e C&alpha;-carboxila permitem verificar a liberdade de rotação característica das cadeias polipeptídicas e as possibilidades dos ângulos de torção &Psi; e &Phi;, visualizando a restrição de rotação da ligação peptídica. Montando um conjunto de aminoácidos é possível construir uma cadeia polipeptídica e, através das ligações de hidrogênio, montar as estruturas secundárias principais (hélice-&alpha; e estruturas &beta;). Duas avaliações preliminares foram realizadas e a avaliação final ocorreu em uma oficina de atividades com 256 professores das áreas de ciências da natureza da rede publica do Estado de SP. Os resultados da avaliação foram extremamente positivos, sendo importante destacar a quantidade e o teor dos comentários elogiosos ao potencial de utilização do material, notadamente, dos professores de biologia e química. O material poderá inclusive auxiliar no preenchimento de lacunas conceituais que existem na formação dos professores e que foram observadas durante as atividades de avaliação. Este conjunto de peças, organizado na forma de um kit: Construindo Estruturas de Aminoácidos e Proteínas, foi submetido à avaliação do MEC e certificado por este órgão, passando a integrar o Guia de Tecnologias Educacionais 2008.<b/> / It was developed a set of plastic pieces that allow the assembly and representation of the most common amino acids, as well as the construction of protein structures. During and after development the material was submitted to several stages of evaluation by teachers (primary and university), graduate and undergraduate students. The first step was the development of models in the computing environment, followed by prototyping of parts. After discussion with the scientific community (presented at the XXXVI Annual Meeting of SBBq in 2007) suggestions were implemented in the computational models. Four thermoplastic injection molds were designed, detailed and constructed under our supervision. Parts representing the structures of amino acids and bonds were produced in large scale and it was started the final process of evaluation. The pieces had good geometric relationships with the structural formulas of amino acids obtained from databases and textbooks. The connections C&alpha;-amine and C&alpha;-arboxyl permit to check the freedom of rotation of the polypeptide chains and the possibility of torsion angles &Phi; and &Psi;, visualizing the restriction of rotation of the peptide bond. Assembling a set of amino acids is possible to build a polypeptide chain and, through hydrogen bonding, to assemble the main secondary structures (&alpha;-helix and &beta;-structures). Two preliminary evaluations were conducted and the final evaluation took place in a workshop with 256 teachers of the fields of natural sciences from public schools of the São Paulo State. The results of the evaluation were extremely positive and it is important to highlight the amount and content of approving comments for the potential of use of the material, especially from biology and chemistry teachers. The material may even assist in filling in conceptual gaps that exist in teacher instruction and that were observed during the evaluation activities. This set of pieces, arranged in the form of a kit: Building Structures of Amino Acids and Proteins, was submitted to MEC and certified by this organization, starting to integrate the Guide of Educational Technology 2008<b/>.

The genetic code of all known organisms is comprised of the 20 proteinogenic amino acids that serve as building blocks on a peptide chain to form a vast array of proteins. Proteins are responsible for virtually every biological process in all organisms; however, the 20 amino acids contain a limited number of functional groups that often leaves much to be desired. The lack of diversity addresses the need to increase the genetic repertoire of living cells to include a variety of amino acids with novel structural, chemical, and physical properties not found in the common 20 amino acids. In order to expand the chemical scope of the genetic code beyond the functionalities that can be directly genetically encoded, unnatural amino acids must be added to the proteome. The ability to incorporate unnatural amino acids (UAAs) into proteins at defined sites has a direct impact on the ability of scientists to study biological processes that are difficult or impossible to address by more classical methods. The UUAs of interest are acylated lysine derivatives (isovaleryl, isobutyryl, and β-hydroxybutyryl) and photocaged histidine. Acylation of histone lysine has been linked to epigenetic regulation of metabolism.1 A means to site-specifically incorporate each acylated lysine derivative would help study the effect of acylated lysine in epigenetic regulation. Likewise, in order to elucidate the role of histidine in specific protein functions, one can replace a critical histidine with a photocaged histidine. Photocaged amino acids are those that possess a photo-cleavable, aromatic caged group. Light-induced protein activation allows for the biological activity of the protein to be spatiotemporally regulated under non-invasive external control.2 The site-specific in vivo incorporation of unnatural amino acids is made possible by amber codon suppression by an orthogonal suppressor aminoacyl-tRNA synthetase (aaRS)/tRNA pair.3 In amber codon suppression the amber stop codon is decoded for an UAA by a suppressor aaRS/tRNA pair. To accept the UAA, the aaRS must be evolved to achieve orthogonal activity with specific UUAs. The pyrrolysyl aaRS/tRNA (PylRS/PylT) pair from M. barkeri and M. mazei was used to construct multiple, large-scale aaRS mutant libraries where critical residues within the active site of PylRS are mutated via site-saturated mutagenesis.4 The libraries were subjected to directed evolution through a series of positive and negative selections to enrich aaRS variants that exclusively bind to acylated lysine derivatives and photocaged histidine as substrates.5 The PylRS selection survivors were screened for UAA activity and identified successful clones underwent a fluorescent activity assay. The active aaRS were used for amber codon suppression to express the respective UAA in ubiquitin and green fluorescent protein constructs.

Sudden changes in nutrient resources are common in the natural environment. Cells are able to adapt and propagate under changing environmental conditions by making adjustments in their cellular processes. These cellular adaptations involve genome-wide transcriptional reprogramming that results in the induction or repression of metabolic pathways. Specific enzymes are then synthesised and activated to maximise the use of the newly available nutrient sources. L-serine is one of the twenty proteinogenic amino acids, and can be synthesised in yeast by the glycolytic and gluconeogenic pathways when growing on fermentable or non-fermentable carbon sources or taken up from the environment when available. L-serine is metabolically linked to glycine and is a predominant donor of one-carbon units in one-carbon metabolism. L-serine is also a source of pyruvate and ammonia and contributes to other cellular processes including the biosynthesis of cysteine and phospholipids. Previous work has shown that yeast cells exhibit transcriptional induction of the one-carbon pathway and the genes involved in the synthesis of purine and methionine after the addition of 10 mM glycine. Here it is shown that addition of 10 mM L-serine did not, however, elicit the same transcriptional response. This is primarily due to differences in the uptake of glycine and L-serine in yeast. High concentrations of extracellular L-serine were required for yeast to show an increase in intracellular L-serine concentration of the magnitude required to trigger a noticeable cellular response. Despite L-serine and glycine being interconvertable via the SHMT isozymes and being a one-carbon donor, the genome-wide transcriptional response exhibited by cells in response to L-serine addition was markedly different to that seen for glycine. The predominant response to an increase in intracellular L-serine was the induction of the general amino acid control system and the CHA1 gene encoding the serine (threonine) dehydratase. Unlike glycine, addition of L-serine triggered only minor induction of the one-carbon pathway. A large portion of intracellular L-serine was converted to pyruvate and ammonia in the mitochondrion as the result of induction of CHA1. The high intracellular concentration of L-serine stimulated the cell to increase the production of oxaloacetate and to increase the biosynthesis of L-aspartate. Transient increases in the intracellular L-glutamate and L-glutamine were also observed after the addition of L-serine. The work presented in this study shows that large increase in the intracellular concentration of amino acid is required to trigger a significant transcriptional response. Yeast cells exhibit different transcriptional and metabolic responses to the addition of L-serine and glycine even though these two amino acids are closely metabolically linked. Addition of L-serine provokes the GAAC response, expression of the CHA1 gene and stimulates the biosynthesis of L-aspartate in yeast whereas addition of glycine induces the one-carbon pathway which leads to the biosynthesis of the purine nucleotides.

Bibliography: leaves 226-261. Investigates nitrogen utilization in some species of rumen bacteria with the object of understanding the role of ammonia versus exogenous amino acids in relation to microbial growth.