IMMUNOCHEMICAL QUANTITATIVE DETERMINATION OF SOME SERUM PROTEINS AND THEIR DIAGNOSTICAL IMPORTANCE

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RELATION BETWEEN VARICELLA AND HERPES ZOSTER

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DYSPROTEINAEMIA IN VIRAL HEPATITIS

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QUANTITATIVE CHANGES IN SOME SERUM PROTEINS IN VIRAL HEPATITIS

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INDIVIDUAL SERUM PROTEIN CONSTELLATIONS IN THE DIAGNOSIS AND PROGNOSIS OF VIRAL HEPATITIS

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UNVEILING NOVEL ASPECTS OF D-AMINO ACID METABOLISM IN THE MODEL BACTERIUM PSEUDOMONAS PUTIDA KT2440

D-amino acids (D-AAs) are the α-carbon enantiomers of L-amino acids (L- AAs), the building blocks of proteins in known organisms. It was largely believed that D-AAs are unnatural and must be toxic to most organisms, as they would compete with the L-counterparts for protein synthesis. Recently, new methods have been developed that allow scientists to chromatographically separate the two AA stereoisomers. Since that time, it has been discovered that D-AAs are vital molecules and they have been detected in many organisms. The work of this dissertation focuses on their place in bacterial metabolism. This specific area was selected due to the abundance of D-AAs in bacteria-rich environments and the knowledge of their part in several processes, such as peptidoglycan synthesis, biofilm disassembly, and sporulation. We focused on the bacterium Pseudomonas putida KT2440 which inhabits the densely populated plant rhizosphere. Due to its versatility and cosmopolitan character, this bacterium has provided an excellent system to study D-AA metabolism. In the first chapter, we have developed a new approach to identify specific genes encoding enzymes acting on D-AAs, collectively known as amino acid racemases. Using this novel method, we identified three amino acid racemases encoded by the genome of P. putida KT2440. All of the enzymes were subsequently cloned and purified to homogeneity, followed by a complete biochemical characterization. The aim of the second chapter was to understand the specific role of the peculiar broad-spectrum amino acid racemase Alr identified in chapter one. After constructing a markerless deletion of the cognate gene, we conducted a variety of phenotypic assays that led to a model for a novel catabolic pathway that involves D-ornithine as an intermediate. The work in chapter three identifies for the first time numerous rhizosphere-dwelling bacteria capable of catabolizing D-AAs. Overall, the work in this dissertation contributes a novel understanding of D-AA catabolism in bacteria and aims to stimulate future efforts in this research area

Aspects of personal data protection in handling cardiac data

Сърдечната недостатъчност като крайна фаза на всички сърдечни заболявания е проблем, за решаването на който е необходимо лекарят да притежава голям обем информация, свързана със състоянието на пациентите. Процесите на консултации, диагностика и лечение са свързани с обработка на специални категории лични данни, която трябва да бъде приведена в съответствие с общия регламент относно защитата на данните на физическите лица, приет на 4 май 2016г и влизащ в сила от 25 Май 2018г. В доклада се изясняват задължителните изисквания, които е необходимо да бъдат изпълнени, и се дават препоръки за приложимите ор-ганизационни и технически мерки за постигане на съответствие с регламента.Heart failure as an end stage of all heart cardiovascular diseases is a problem for which a physician needs to have a large amount of information related to the condition of patients. Consultation, diagnosis and treatment processes are related to the processing of special categories of personal data, which must be brought into line with the General Data Protection Regulation of Natural Persons adopted on May 4, 2016 and in force since May 25, 2018. The report clarifies the mandatory requirements that need to be met and provides recommendations on the applicable organizational and technical measures to comply with the Regulation

A Broad Spectrum Racemase in \u3cem\u3ePseudomonas putida\u3c/em\u3e KT2440 Plays a Key Role in Amino Acid Catabolism

The broad-spectrum amino acid racemase (Alr) of Pseudomonas putida KT2440 preferentially interconverts the L- and D-stereoisomers of Lys and Arg. Despite conservation of broad-spectrum racemases among bacteria, little is known regarding their physiological role. Here we explore potential functional roles for Alr in P. putida KT2440. We demonstrate through cellular fractionation that Alr enzymatic activity is found in the periplasm, consistent with its putative periplasm targeting sequence. Specific activity of Alr is highest during exponential growth, and this activity corresponds with an increased accumulation of D-Lys in the growth medium. An alr gene knockout strain (Δalr) was generated and used to assess potential roles for the alr gene in peptidoglycan structure, producing soluble signaling compounds, and amino acid metabolism. The stationary phase peptidoglycan structure did not differ between wild-type and Δalr strains, indicating that products resulting from Alr activity are not incorporated into peptidoglycan under these conditions. RNA-seq was used to assess differences in the transcriptome between the wild-type and Δalr strains. Genes undergoing differential expression were limited to those involved in amino acid metabolism. The Δalr strain exhibited a limited capacity for catabolism of L-Lys and L-Arg as the sole source of carbon and nitrogen. This is consistent with a predicted role for Alr in catabolism of L-Lys by virtue of its ability to convert L-Lys to D-Lys, which is further catabolized through the L-pipecolate pathway. The metabolic profiles here also implicate Alr in catabolism of L-Arg, although the pathway by which D-Arg is further catabolized is not clear at this time. Overall, data presented here describe the primary role of Alr as important for basic amino acid metabolism

NOTES ON SOME SERUM PROTEIN CHANGES IN VIRAL HEPATITIS - BIOCHEMICAL ASPECTS

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Amino Acid Racemase Enzyme Assays

Amino acid racemases are enzymes that invert the α-carbon stereochemistry of amino acids (AAs), interconverting amino acids between their L- and D-enantiomers in a reversible reaction. In bacteria, they are known to have catabolic physiological functions but are also involved in the synthesis of many D-AAs, including D-glutamate and D-alanine, which are necessary components of the peptidoglycan layer of the bacterial cell wall. As such, amino acid racemases represent significant targets for the development of bactericidal compounds. Amino acid racemases are also regarded by the biotechnological industry as important catalysts for the production of economically relevant D-AAs. Here, we provide a detailed protocol using high performance liquid chromatography (HPLC) and 1-fluoro-2,4-dinitrophenyl-5-L-alanine amide (FDAA, also Marfey’s reagent) for the characterization of novel amino acid racemases. The protocol described here was designed to obtain accurate kinetic parameters (kcat, KM values). Enzyme concentrations and reaction times were optimized so as to minimize the reverse reaction, which can confound results when measuring racemase reactions