NÁLEZKOVÁ, Monika, Lukáš ŽÍDEK and Vladimír SKLENÁŘ. NMR Study of Protein CD69. In Strukturni biofyzika makromolekul. Czech Republic: MU, 2005, p. 13.
Other formats:   BibTeX LaTeX RIS
Basic information
Original name NMR Study of Protein CD69
Name in Czech Stidium proteinu CD69 pomoci NMR
Authors NÁLEZKOVÁ, Monika, Lukáš ŽÍDEK and Vladimír SKLENÁŘ.
Edition Czech Republic, Strukturni biofyzika makromolekul, p. 13-13, 2005.
Publisher MU
Other information
Original language English
Type of outcome Proceedings paper
Field of Study 10610 Biophysics
Country of publisher Czech Republic
Confidentiality degree is not subject to a state or trade secret
Organization unit Faculty of Science
Keywords in English NMR CD69 lectin-type proteins NK cell glycoproteins
Changed by Changed by: prof. Mgr. Lukáš Žídek, Ph.D., učo 38990. Changed: 9/4/2010 14:03.
Abstract
The lectin-type proteins, like CD69, are transmembrane, disulphide-linked glycoproteins of natural killer (NK) cells. NK cells play an important role in the immune response to cell-surface displayed antigens, such as those found on virus infected cells, tumor cells, etc. Several oligosacharides were suggested to be natural ligands of CD69 receptor and NMR method was found convenient for observation of CD69 sacharide interaction. NMR is noninvasive method, able to observe molecules in environment close to physiological conditions. Unfortunately, NMR is not very sensible and big amount of sample (300microl of 0.2mM at least), usually enriched by nuclei 15N and/or 13C, is needed. Preparation of well folded highly concentrated protein sample is thus crucial. At the beggining of this study, the construct of approximatelly 130 amino acid long extracelular part of human CD69 was prepared by the group of Karel Bezouška, Department of Biochemistry, Faculty of Science, Prague. NMR measurement of the construct showed that protein is totally unfolded. A construct of homologue of rat CD69 was then delivered. Basic 2D NMR experiment found partially structured protein. In the summer 2005, the problem with folding of human CD69 was solved by our collaborators and experiments aimed at ligand binding were started. As binding of sacharides to CD69 receptor is supposed to be dependent on presence of Ca2+, titration experiments with CaCl2 were performed, followed by titration with GlcNAc and pentaantenary oligosacharide. It was posible to observe several small changes of chemical shift in NMR spectra correlating chemical shifts of hydrogen and nitrogen. In order to study protein-ligand interactions in more details it was decided to prepare double (15N, 13C) labelled sample, and to assign side chain nuclei of the protein. The double-labelled sample has been obtained recently and assignment is currently in progress.
Abstract (in Czech)
The lectin-type proteins, like CD69, are transmembrane, disulphide-linked glycoproteins of natural killer (NK) cells. NK cells play an important role in the immune response to cell-surface displayed antigens, such as those found on virus infected cells, tumor cells, etc. Several oligosacharides were suggested to be natural ligands of CD69 receptor and NMR method was found convenient for observation of CD69 sacharide interaction. NMR is noninvasive method, able to observe molecules in environment close to physiological conditions. Unfortunately, NMR is not very sensible and big amount of sample (300microl of 0.2mM at least), usually enriched by nuclei 15N and/or 13C, is needed. Preparation of well folded highly concentrated protein sample is thus crucial. At the beggining of this study, the construct of approximatelly 130 amino acid long extracelular part of human CD69 was prepared by the group of Karel Bezouška, Department of Biochemistry, Faculty of Science, Prague. NMR measurement of the construct showed that protein is totally unfolded. A construct of homologue of rat CD69 was then delivered. Basic 2D NMR experiment found partially structured protein. In the summer 2005, the problem with folding of human CD69 was solved by our collaborators and experiments aimed at ligand binding were started. As binding of sacharides to CD69 receptor is supposed to be dependent on presence of Ca2+, titration experiments with CaCl2 were performed, followed by titration with GlcNAc and pentaantenary oligosacharide. It was posible to observe several small changes of chemical shift in NMR spectra correlating chemical shifts of hydrogen and nitrogen. In order to study protein-ligand interactions in more details it was decided to prepare double (15N, 13C) labelled sample, and to assign side chain nuclei of the protein. The double-labelled sample has been obtained recently and assignment is currently in progress.
Links
LC06030, research and development projectName: Biomolekulární centrum
Investor: Ministry of Education, Youth and Sports of the CR, Biomolecular centre
PrintDisplayed: 27/4/2024 20:42