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Modification of glutathione S-transferase by selected glycating agents
Průchová, Zuzana ; Dršata, Jaroslav (advisor) ; Trnková, Lucie (referee)
Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Biochemical Sciences Candidate: Zuzana Průchová Supervisor: Prof. MUDr. Jaroslav Dršata, CSc. Title of diploma thesis: Modification of Glutathione S-transferase by selected glycating agents Glycation is a non-enzymatic reaction, which leads to the binding of carbonyl compounds to free amino groups of amino acids, peptides, proteins or other biomolecules. This process is closely related to impaired protein function in diabetes mellitus, during aging, and in neurodegenerative diseases. The main purpose of this thesis was to monitor the impact of several glycating agents on the properties of the enzyme glutathione S-transeferase (GST, EC 2.5.1.18) which plays a key role in the detoxification of many exogenous as well as endogenous compounds. Methylglyoxal (0.5-2 mM), glyceraldehyde (0.5-10 mM), glucose (50 mM), and fructose (50 mM) were selected as glycating agents. The course of protein glycation was evaluated by following methods: GST catalytic activity assessment, determination of primary amino groups, assessment of AGEs formation using fluorescence and Western blotting, generation of high molecular cross-links and aggregates by denaturationg polyacrylamide gel electrophoresis (SDS-PAGE), changes in GST molecular...
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Tumor markers
Karmazínová, Simona ; Dršata, Jaroslav (advisor) ; Skálová, Lenka (referee)
Tumorous diseases are as old as life itself. They pose a serious social problem and everybody has encountered them at some time. Due to the polluted environment and modern way of living, tumorous diseases are becoming increasingly responsible for human deaths. Tumorous diseases can appear in any multicellular organism in some form. Traces of them have been identified in prehistoric people as well as in Egyptian mummies. The widespread word CANCER has its origin in ancient Greece. The first reports of laboratory evidence of tumorous diseases were published in scientific publications as early as the mid-19th century. This was the time of the onset of clinical biochemistry as a branch of science and medicine. Laboratory techniques did not exist then and testing was based on simple physico-chemical reactions. It was only in the second half of the 20th century that the study and examination of tumour markers enjoyed rapid development. Currently, the most common tumour can be determined by nearly any biochemical laboratory. The subject of my paper, "Tumour markers" is very broad - actually, each chapter would deserve to be an independent subject - and still, my thesis is far from a detailed analysis of each individual marker and methods of its determination. I aimed at providing a brief overview of the...
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Influence of pyridoxalisonicotinoyl hydrazone on spectral properties of alanine aminotransferase
Vančurová, Kamila ; Dršata, Jaroslav (advisor) ; Skálová, Lenka (referee)
The aim of this study was to examine the influence of pyridoxalisonicotinoylhydrazone (PIH) on alanine aminotransferase (EC 2. 6. 1. 2.). Alanine aminotransferase belongs to the group of pyridoxal enzymes. Pyridoxal-5'-phosphate is a prosthetic group of these enzymes. There is a similarity to the structure of pyridoxal and isoniazide in the formula of pyridoxalisonicotinoylhydrazone. PIH has been investigated for its potential iron-chelating activity. There are also studies about PIH's protective effect against cardiotoxicity caused by the treatment with anthracyclines. Cardiotoxicity is the main harmful side effect of the anthracycline medication. There is a part which is similar to pyridoxalphosphate in the structure of PIH. That's why the influence of PIH on spectral properties of pyridoxal enzymes was studied. For this spectral study we chose transaminases as suitable pyridoxal enzymes. During the catalyzed reaction they change their absorption spectra. Most common transaminase aspartate aminotransferase (AST) was not optimal for this study. AST absorbs in the same region where PIH does. Alanine aminotransferase absorbs in the spectrum separately and therefore it was convenient for this study. UV-VIS spectra and circular dichroism (CD) spectra were measured. The ability of providing CD spectra is the...
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Biological activity of potential antioxidants in L-6 rat myoblasts
Drahošová, Vlasta ; Boušová, Iva (advisor) ; Dršata, Jaroslav (referee)
Charles University in Prague University Roma Tre Faculty of Pharmacy in Hradec Králové Department of Sciences Department of Biochemical Sciences Laboratory of Physiology Candidate: Vlasta Drahošová Supervisor: Doc. PharmDr. Iva Boušová, Ph.D. Prof. Sandra Incerpi Title of diploma thesis: Biological activity of potential antioxidants in L-6 rat myoblasts This diploma thesis is based on the results of research conducted at the Department of Sciences, University of Roma Tre in Rome under the supervision of Professor Sandra Incerpi. Team of the Laboratory of physiology has been involved in the research of various natural substances with antioxidant and cytostatic properties for many years. Antioxidants are able to eliminate reactive oxygen species (ROS), which exert irreplaceable physiological effects in the body, but they are also a cause of certain diseases progression and aging due to the damage of various biomolecules. In this work, I studied the antioxidant activity of six synthetic derivatives of 4-methylcoumarin (AB-3, AB-4, AB-7, AB-8, AB-11, AB-12), which were synthesized and characterized by prof. Virinder Parmar (University of Delhi, India). Study was conducted in the L-6 rat myoblasts cell line (determination of intracellular ROS using dichlorofluorescein assay, cell proliferation assay)...
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Expression of cytochrome P450 2C9 in liver tumors
Štěpařová, Andrea ; Dršata, Jaroslav (advisor) ; Skálová, Lenka (referee)
2 ABSTRACT Bc. Andrea Štěpařová Expression of cytochrome P450 2C9 in hepatic tumors. Diploma thesis Charles University in Prague, Faculty of Pharmacy in Hradec Králové Healthcare bioanalytics - Specialist in Laboratory Methods Background: Liver cancer is a growing health problem worldwide. The most common primary hepatic malignancy is hepatocellular carcinoma (HCC). It is a multifactorial disease arising from multiple genetic and non-genetic alterations, most importantly viral infections and toxic injury (e.g. alcohol abuse). Cytochromes P450 (CYPs) participate on a plethora of metabolic processes in human body. Several CYPs play a role in tumorigenesis as well. This theses covers the topics of hepatic malignancies and regulation of cytochrome P450 expression to evaluate the possible relationship to the hepatocarcinogenesis and pharmacotherapy of HCC. In CYP2C9, which is one of the most important drug-metabolizing CYP forms, gene expression analysis was performed in HCC tumor and surrounding tissue, also including CYP2C9 major regulators - nuclear receptors PXR (pregnane-X-receptor) and CAR (constitutive androstane receptor). Methods: Samples of hepatocellular carcinoma and surrounding non-cancerous tissue from 12 patients were obtained from the biobank of Masaryk Memorial Cancer Institute, Brno, Czech...
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