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Recombinant expression and studies on DCL-1, receptor of dendritic cells
Pospíšilová, Eliška ; Bezouška, Karel (advisor) ; Novák, Petr (referee)
Recombinant expression and studies on DCL-1, receptor of dendritic cells Eliška Pospíšilová Charles University in Prague, Faculty of Science, Department of Biochemistry ABSTRACT Dendritic cells, which could be found in large numbers in many tissues, exprime C-type lectine receptor DCL-1 (in CD nomenclature CD302) on their surface. This molecule belongs to the class one transmembrane proteins. N-terminal sequence a C-type lectin-like domain (CTLD) it has on the extracellular side. Whereas the C-terminus, which contains potential phosphorylation site and signals for intracellular transport, is in the cytosol. As the sequence of this protein is highly conserved through all mammalian species, it is likely to play an important role in the immune system. Despite this fact, the DCL-1 molecule is still poorly investigated. This work deals with the extracellular part - especcially with the CTLD containing part - of the DCL-1 receptor. To learn more about this molecule, we attempted its recombinant expression in bacteria. We used pET-30a(+) based vector pDCL1E, and bacterial cells E. coli BL21 (DE3) Gold, which produced the target protein in the form of inclusion bodies into the cytoplasm. Therefore conditions for in vitro renaturation were optimalized, and it was prooved that the protein had native conformation...
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Recombinant expression and studies on DCL-1, receptor of dendritic cells
Pospíšilová, Eliška ; Bezouška, Karel (advisor) ; Novák, Petr (referee)
Recombinant expression and studies on DCL-1, receptor of dendritic cells Eliška Pospíšilová Charles University in Prague, Faculty of Science, Department of Biochemistry ABSTRACT Dendritic cells, which could be found in large numbers in many tissues, exprime C-type lectine receptor DCL-1 (in CD nomenclature CD302) on their surface. This molecule belongs to the class one transmembrane proteins. N-terminal sequence a C-type lectin-like domain (CTLD) it has on the extracellular side. Whereas the C-terminus, which contains potential phosphorylation site and signals for intracellular transport, is in the cytosol. As the sequence of this protein is highly conserved through all mammalian species, it is likely to play an important role in the immune system. Despite this fact, the DCL-1 molecule is still poorly investigated. This work deals with the extracellular part - especcially with the CTLD containing part - of the DCL-1 receptor. To learn more about this molecule, we attempted its recombinant expression in bacteria. We used pET-30a(+) based vector pDCL1E, and bacterial cells E. coli BL21 (DE3) Gold, which produced the target protein in the form of inclusion bodies into the cytoplasm. Therefore conditions for in vitro renaturation were optimalized, and it was prooved that the protein had native conformation...
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