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Preparation of strategy for studying interaction of human constitutive androstane receptor with steroid receptor coactivator-1
Šachová, Ivana ; Martin, Jan (advisor) ; Mladěnka, Přemysl (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Pharmacognosy Candidate: Ivana Šachová Supervisors: PharmDr. Jan Martin, PhD., Dipl. Ing. Mgr. Ferdinand Molnár, PhD. Title of diploma thesis: Preparation of strategy for studying interaction of human constitutive androstane receptor with steroid receptor coactivator-1 The aim of this study was to express wild type and mutated synthetic human SRC1 central domains in E. coli BL21 DE3 cells. Expressed recombinant proteins were purified afterwards and used for investigation of SRC1-CAR protein interaction using pull down experiments. The constitutive androstane receptor (CAR), a nuclear receptor (NR), is a transcriptional regulator, which influences the expression of various proteins, such as enzymes of biotransformation and transporters important for metabolism of both endogenous and exogenous compounds. CAR can be fully active only in presence of coactivators, such as steroid receptor coactivator-1 (SRC1). The most important fragment of SRC1 for binding to NRs is the central domain, made of three α-helical Leu-X-X-Leu-Leu (LXXLL) L1, L2, L3 motifs, in where L is a leucine and X represents any amino acid. hSRC1, as well as hCAR, can be successfully produced by expression of cDNA of interest in E. coli strains. In case of SRC1,...
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Preparation of strategy for studying interaction of human constitutive androstane receptor with steroid receptor coactivator-1
Šachová, Ivana ; Martin, Jan (advisor) ; Mladěnka, Přemysl (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Pharmacognosy Candidate: Ivana Šachová Supervisors: PharmDr. Jan Martin, PhD., Dipl. Ing. Mgr. Ferdinand Molnár, PhD. Title of diploma thesis: Preparation of strategy for studying interaction of human constitutive androstane receptor with steroid receptor coactivator-1 The aim of this study was to express wild type and mutated synthetic human SRC1 central domains in E. coli BL21 DE3 cells. Expressed recombinant proteins were purified afterwards and used for investigation of SRC1-CAR protein interaction using pull down experiments. The constitutive androstane receptor (CAR), a nuclear receptor (NR), is a transcriptional regulator, which influences the expression of various proteins, such as enzymes of biotransformation and transporters important for metabolism of both endogenous and exogenous compounds. CAR can be fully active only in presence of coactivators, such as steroid receptor coactivator-1 (SRC1). The most important fragment of SRC1 for binding to NRs is the central domain, made of three α-helical Leu-X-X-Leu-Leu (LXXLL) L1, L2, L3 motifs, in where L is a leucine and X represents any amino acid. hSRC1, as well as hCAR, can be successfully produced by expression of cDNA of interest in E. coli strains. In case of SRC1,...
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Preparation of strategy for studying interaction of human constitutive androstane receptor with steroid receptor coactivator-1
Šachová, Ivana ; Martin, Jan (advisor) ; Mladěnka, Přemysl (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Pharmacognosy Candidate: Ivana Šachová Supervisors: PharmDr. Jan Martin, PhD., Dipl. Ing. Mgr. Ferdinand Molnár, PhD. Title of diploma thesis: Preparation of strategy for studying interaction of human constitutive androstane receptor with steroid receptor coactivator-1 The aim of this study was to express wild type and mutated synthetic human SRC1 central domains in E. coli BL21 DE3 cells. Expressed recombinant proteins were purified afterwards and used for investigation of SRC1-CAR protein interaction using pull down experiments. The constitutive androstane receptor (CAR), a nuclear receptor (NR), is a transcriptional regulator, which influences the expression of various proteins, such as enzymes of biotransformation and transporters important for metabolism of both endogenous and exogenous compounds. CAR can be fully active only in presence of coactivators, such as steroid receptor coactivator-1 (SRC1). The most important fragment of SRC1 for binding to NRs is the central domain, made of three α-helical Leu-X-X-Leu-Leu (LXXLL) L1, L2, L3 motifs, in where L is a leucine and X represents any amino acid. hSRC1, as well as hCAR, can be successfully produced by expression of cDNA of interest in E. coli strains. In case of SRC1,...
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