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dc.contributor.author
von Stetten, David
dc.contributor.author
Seibeck, Sven
dc.contributor.author
Michael, Norbert
dc.contributor.author
Scheerer, Patrick
dc.contributor.author
Mroginski, Maria Andrea
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dc.contributor.author
Murgida, Daniel Horacio
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dc.contributor.author
Krauss, Norbert
dc.contributor.author
Heyn, Maarten P.
dc.contributor.author
Hildebrandt, Peter
dc.contributor.author
Borucki, Berthold
dc.contributor.author
Lamparter, Tilman
dc.date.available
2020-04-28T22:37:08Z
dc.date.issued
2007-01-19
dc.identifier.citation
von Stetten, David; Seibeck, Sven; Michael, Norbert; Scheerer, Patrick; Mroginski, Maria Andrea; et al.; Highly conserved residues Asp-197 and His-250 in Agp1 phytochrome control the proton affinity of the chromophore and Pfr formation; American Society for Biochemistry and Molecular Biology; Journal of Biological Chemistry (online); 282; 3; 19-1-2007; 2116-2123
dc.identifier.issn
0021-9258
dc.identifier.uri
http://hdl.handle.net/11336/103859
dc.description.abstract
The mutants H250A and D197A of Agp1 phytochrome from Agrobacterium tumefaciens were prepared and investigated by different spectroscopic and biochemical methods. Asp-197 and His-250 are highly conserved amino acids and are part of the hydrogen-bonding network that involves the chromophore. Both substitutions cause a destabilization of the protonated chromophore in the Pr state as revealed by resonance Raman and UV-visible absorption spectroscopy. Titration experiments demonstrate a lowering of the pK(a) from 11.1 ( wild type) to 8.8 in H250A and 7.2 in D197A. Photoconversion of the mutants does not lead to the Pfr state. H250A is arrested in a meta-Rc-like state in which the chromophore is deprotonated. For H250A and the wild-type protein, deprotonation of the chromophore in meta-Rc is coupled to the release of a proton to the external medium, whereas the subsequent proton re-uptake, linked to the formation of the Pfr state in the wild- type protein, is not observed for H250A. No transient proton exchange with the external medium occurs in D197A, suggesting that Asp-197 may be the proton release group. Both mutants do not undergo the photoinduced protein structural changes that in the wild- type protein are detectable by size exclusion chromatography. These conformational changes are, therefore, attributed to the meta-Rc -> Pfr transition and most likely coupled to the transient proton re- uptake. The present results demonstrate that Asp-197 and His-250 are essential for stabilizing the protonated chromophore structure in the parent Pr state, which is required for the primary photochemical process, and for the complete photo-induced conversion to the Pfr state.
dc.format
application/pdf
dc.language.iso
eng
dc.publisher
American Society for Biochemistry and Molecular Biology
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dc.rights
info:eu-repo/semantics/openAccess
dc.rights.uri
https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
dc.subject
PHYTOCHROME
dc.subject
PHOTORECEPTORS
dc.subject
FT-RAMAN
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DFT CALCULATIONS
dc.subject
CLONING, MOLECULAR
dc.subject
MUTATION
dc.subject.classification
Físico-Química, Ciencia de los Polímeros, Electroquímica
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dc.subject.classification
Ciencias Químicas
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dc.subject.classification
CIENCIAS NATURALES Y EXACTAS
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dc.title
Highly conserved residues Asp-197 and His-250 in Agp1 phytochrome control the proton affinity of the chromophore and Pfr formation
dc.type
info:eu-repo/semantics/article
dc.type
info:ar-repo/semantics/artículo
dc.type
info:eu-repo/semantics/publishedVersion
dc.date.updated
2020-04-22T15:40:50Z
dc.journal.volume
282
dc.journal.number
3
dc.journal.pagination
2116-2123
dc.journal.pais
Estados Unidos
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dc.journal.ciudad
Bethesda
dc.description.fil
Fil: von Stetten, David. Technische Universität Berlin; Alemania
dc.description.fil
Fil: Seibeck, Sven. Freie Universität Berlin.; Alemania
dc.description.fil
Fil: Michael, Norbert. Freie Universität Berlin.; Alemania
dc.description.fil
Fil: Scheerer, Patrick. Charité Universitätsmedizin Berlin; Alemania
dc.description.fil
Fil: Mroginski, Maria Andrea. Technische Universität Berlin; Alemania
dc.description.fil
Fil: Murgida, Daniel Horacio. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Química, Física de los Materiales, Medioambiente y Energía. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Química, Física de los Materiales, Medioambiente y Energía; Argentina. Technische Universität Berlin; Alemania
dc.description.fil
Fil: Krauss, Norbert. Freie Universität Berlin.; Alemania
dc.description.fil
Fil: Heyn, Maarten P.. Charité Universitätsmedizin Berlin; Alemania
dc.description.fil
Fil: Hildebrandt, Peter. Technische Universität Berlin; Alemania
dc.description.fil
Fil: Borucki, Berthold. Freie Universität Berlin.; Alemania
dc.description.fil
Fil: Lamparter, Tilman. Freie Universität Berlin.; Alemania
dc.journal.title
Journal of Biological Chemistry (online)
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dc.relation.alternativeid
info:eu-repo/semantics/altIdentifier/url/https://www.jbc.org/content/282/3/2116
dc.relation.alternativeid
info:eu-repo/semantics/altIdentifier/doi/http://dx.doi.org/10.1074/jbc.M608878200
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