Chromophore structure in the photocycle of the cyanobacterial phytochrome Cph1.
The chromophore conformations of the red and far red light induced product states "Pfr" and "Pr" of the N-terminal photoreceptor domain Cph1-N515 from Synechocystis 6803 have been investigated by NMR spectroscopy, using specific 13C isotope substitutions in the chromophore. 13C-N...
| Main Authors: | , , , , |
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| Format: | Journal article |
| Language: | English |
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2006
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| _version_ | 1797061091387244544 |
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| author | van Thor, J Mackeen, M Kuprov, I Dwek, R Wormald, M |
| author_facet | van Thor, J Mackeen, M Kuprov, I Dwek, R Wormald, M |
| author_sort | van Thor, J |
| collection | OXFORD |
| description | The chromophore conformations of the red and far red light induced product states "Pfr" and "Pr" of the N-terminal photoreceptor domain Cph1-N515 from Synechocystis 6803 have been investigated by NMR spectroscopy, using specific 13C isotope substitutions in the chromophore. 13C-NMR spectroscopy in the Pfr and Pr states indicated reversible chemical shift differences predominantly of the C(4) carbon in ring A of the phycocyanobilin chromophore, in contrast to differences of C15 and C5, which were much less pronounced. Ab initio calculations of the isotropic shielding and optical transition energies identify a region for C4-C5-C6-N2 dihedral angle changes where deshielding of C4 is correlated with red-shifted absorption. These could occur during thermal reactions on microsecond and millisecond timescales after excitation of Pr which are associated with red-shifted absorption. A reaction pathway involving a hula-twist at C5 could satisfy the observed NMR and visible absorption changes. Alternatively, C15 Z-E photoisomerization, although expected to lead to a small change of the chemical shift of C15, in addition to changes of the C4-C5-C6-N2 dihedral angle could be consistent with visible absorption changes and the chemical shift difference at C4. NMR spectroscopy of a 13C-labeled chromopeptide provided indication for broadening due to conformational exchange reactions in the intact photoreceptor domain, which is more pronounced for the C- and D-rings of the chromophore. This broadening was also evident in the F2 hydrogen dimension from heteronuclear 1H-13C HSQC spectroscopy, which did not detect resonances for the 13C5-H, 13C10-H, and 13C15-H hydrogen atoms whereas strong signals were detected for the (13)C-labeled chromopeptide. The most pronounced 13C-chemical shift difference between chromopeptide and intact receptor domain was that of the 13C4-resonance, which could be consistent with an increased conformational energy of the C4-C5-C6-N2 dihedral angle in the intact protein in the Pr state. Nuclear Overhauser effect spectroscopy experiments of the 13C-labeled chromopeptide, where chromophore-protein interactions are expected to be reduced, were consistent with a ZZZssa conformation, which has also been found for the biliverdin chromophore in the x-ray structure of a fragment of Deinococcus radiodurans bacteriophytochrome in the Pr form. |
| first_indexed | 2024-03-06T20:26:08Z |
| format | Journal article |
| id | oxford-uuid:2f73c118-2eaa-476c-9d6a-12c49db77c75 |
| institution | University of Oxford |
| language | English |
| last_indexed | 2024-03-06T20:26:08Z |
| publishDate | 2006 |
| record_format | dspace |
| spelling | oxford-uuid:2f73c118-2eaa-476c-9d6a-12c49db77c752022-03-26T12:55:26ZChromophore structure in the photocycle of the cyanobacterial phytochrome Cph1.Journal articlehttp://purl.org/coar/resource_type/c_dcae04bcuuid:2f73c118-2eaa-476c-9d6a-12c49db77c75EnglishSymplectic Elements at Oxford2006van Thor, JMackeen, MKuprov, IDwek, RWormald, MThe chromophore conformations of the red and far red light induced product states "Pfr" and "Pr" of the N-terminal photoreceptor domain Cph1-N515 from Synechocystis 6803 have been investigated by NMR spectroscopy, using specific 13C isotope substitutions in the chromophore. 13C-NMR spectroscopy in the Pfr and Pr states indicated reversible chemical shift differences predominantly of the C(4) carbon in ring A of the phycocyanobilin chromophore, in contrast to differences of C15 and C5, which were much less pronounced. Ab initio calculations of the isotropic shielding and optical transition energies identify a region for C4-C5-C6-N2 dihedral angle changes where deshielding of C4 is correlated with red-shifted absorption. These could occur during thermal reactions on microsecond and millisecond timescales after excitation of Pr which are associated with red-shifted absorption. A reaction pathway involving a hula-twist at C5 could satisfy the observed NMR and visible absorption changes. Alternatively, C15 Z-E photoisomerization, although expected to lead to a small change of the chemical shift of C15, in addition to changes of the C4-C5-C6-N2 dihedral angle could be consistent with visible absorption changes and the chemical shift difference at C4. NMR spectroscopy of a 13C-labeled chromopeptide provided indication for broadening due to conformational exchange reactions in the intact photoreceptor domain, which is more pronounced for the C- and D-rings of the chromophore. This broadening was also evident in the F2 hydrogen dimension from heteronuclear 1H-13C HSQC spectroscopy, which did not detect resonances for the 13C5-H, 13C10-H, and 13C15-H hydrogen atoms whereas strong signals were detected for the (13)C-labeled chromopeptide. The most pronounced 13C-chemical shift difference between chromopeptide and intact receptor domain was that of the 13C4-resonance, which could be consistent with an increased conformational energy of the C4-C5-C6-N2 dihedral angle in the intact protein in the Pr state. Nuclear Overhauser effect spectroscopy experiments of the 13C-labeled chromopeptide, where chromophore-protein interactions are expected to be reduced, were consistent with a ZZZssa conformation, which has also been found for the biliverdin chromophore in the x-ray structure of a fragment of Deinococcus radiodurans bacteriophytochrome in the Pr form. |
| spellingShingle | van Thor, J Mackeen, M Kuprov, I Dwek, R Wormald, M Chromophore structure in the photocycle of the cyanobacterial phytochrome Cph1. |
| title | Chromophore structure in the photocycle of the cyanobacterial phytochrome Cph1. |
| title_full | Chromophore structure in the photocycle of the cyanobacterial phytochrome Cph1. |
| title_fullStr | Chromophore structure in the photocycle of the cyanobacterial phytochrome Cph1. |
| title_full_unstemmed | Chromophore structure in the photocycle of the cyanobacterial phytochrome Cph1. |
| title_short | Chromophore structure in the photocycle of the cyanobacterial phytochrome Cph1. |
| title_sort | chromophore structure in the photocycle of the cyanobacterial phytochrome cph1 |
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