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001     1302
005     20200610184538.0
024 7 _ |2 Handle
|a 2128/13952
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088 1 _ |a Juel-4272
088 _ _ |a Juel-4272
|2 JUEL
100 1 _ |0 P:(DE-Juel1)VDB57288
|a von der Hocht, Iris
|b 0
|e Coresponding author
|u FZJ
245 _ _ |a Fluorescence Spectroscopy of Recoverin Function and Conformation
260 _ _ |a Jülich
|b Forschungszentrum Jülich GmbH Zentralbibliothek, Verlag
|c 2008
300 _ _ |a 119 p.
336 7 _ |0 PUB:(DE-HGF)11
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|a Dissertation / PhD Thesis
336 7 _ |0 PUB:(DE-HGF)3
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|a Book
336 7 _ |0 2
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|a Thesis
336 7 _ |2 DRIVER
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336 7 _ |2 BibTeX
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490 0 _ |0 PERI:(DE-600)2414853-2
|8 32540
|a Berichte des Forschungszentrums Jülich
|v 4272
|x 0944-2952
500 _ _ |a Record converted from VDB: 12.11.2012
502 _ _ |a Köln, Univ., Diss., 2008
|b Dr. (Univ.)
|c Univ. Köln
|d 2008
520 _ _ |a Recoverin is a neuronal calcium sensor protein (NCS-protein) from the vertebrate photoreceptor which is involved in light adaptation. Recoverin changes its conformation upon sequential binding of two calcium ions. This conformational change induces the extrusion of a covalently attached myristoyl residue from a hydrophobic binding pocket which enables Recoverin to interact with lipid membranes (calcium-myristoyl switch). In this thesis, I report on my investigation of Recoverin’s calcium myristoyl switch, using a recently developed modification of fluorescence correlation spectroscopy (FCS) that is called dual-focus FCS (2fFCS). This method allows for measuring absolute diffusion coefficients with an accuracy of better than a few percent. Recoverin, and the Recoverin mutants E85Q and E121Q which bind only one or no Ca$^{2+}$, respectively, were labeled with the fluorescent dye Alexa647. Differences in the hydrodynamic radius due to conformational changes of Recoverin and its mutants upon calcium binding were monitored by measuring the diffusion coefficient of these molecules as a function of free calcium concentration. The calcium dependent interaction of Recoverin with lipid membranes was measured in solutions of small unilamellar vesicles (SUVs) of different lipid composition. Again, diffusion measurements were used to determine the fraction of free and lipid-bound Recoverin using the strongly different diffusion coefficients of both fractions. To account for the fluorescence brightness difference of the dye label when in solution and close to a lipid membrane, a new three-photon correlation analysis was developed and tested.
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655 _ 7 |a Hochschulschrift
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856 4 _ |u https://juser.fz-juelich.de/record/1302/files/J%C3%BCl_4272_Hocht.pdf
|y OpenAccess
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914 1 _ |y 2008
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