Effect of chromophore exchange on the resonance Raman spectra of recombinant phytochromes

Kneip, Christa; Mozley, David; Hildebrandt, Peter; Gärtner, Wolfgang; Braslavsky, Silvia E.; Schaffner, Kurt

doi:10.1016/s0014-5793(97)00969-1

Cited by 32 publications

(47 citation statements)

References 19 publications

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“…As shown by vibrational techniques, the conformations of the bilin chromophore in the Pr and Pfr states in native plant phytochrome, a dimer of two 124 kDa units (phyA124), resemble that in the photosensory modules phyA65 of oat phytochrome (28,29) and Cph1 of Synechocystis (30). Furthermore, our NMR data add clear evidence that the chromophore and its interactions with the protein are conserved in both states throughout the Cph1/plant phytochrome family.…”

supporting

confidence: 59%

Light-induced chromophore activity and signal transduction in phytochromes observed by ¹³ C and ¹⁵ N magic-angle spinning NMR

Rohmer

Lang

Hughes

et al. 2008

Proc. Natl. Acad. Sci. U.S.A.

144

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Both thermally stable states of phytochrome, Pr and Pfr, have been studied by 13 C and 15 N cross-polarization (CP) magic-angle spinning (MAS) NMR using cyanobacterial (Cph1) and plant (phyA) phytochrome sensory modules containing uniformly 13 C-and 15 N-labeled bilin chromophores. Two-dimensional homo-and heteronuclear experiments allowed most of the 13 C chemical shifts to be assigned in both states. Chemical shift differences reflect changes of the electronic structure of the cofactor at the atomic level as well as its interactions with the chromophore-binding pocket. The chromophore in cyanobacterial and plant phytochromes shows very similar features in the respective Pr and Pfr states. The data are interpreted in terms of a strengthened hydrogen bond at the ring D carbonyl. The red shift in the Pfr state is explained by the increasing length of the conjugation network beyond ring C including the entire ring D. Enhanced conjugation within the -system stabilizes the more tensed chromophore in the Pfr state. Concomitant changes at the ring C propionate carboxylate and the ring D carbonyl are explained by a loss of hydrogen bonding to Cph1-His-290 and transmittance of conformational changes to the ring C propionate via a water network. These and other conformational changes may lead to modified surface interactions, e.g., along the tongue region contacting the bilin chromophore.photomorphogenesis ͉ photoreceptors ͉ chromophore-protein interaction ͉ phycocyanobilin ͉ solid-state NMR

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supporting

confidence: 59%

Light-induced chromophore activity and signal transduction in phytochromes observed by ¹³ C and ¹⁵ N magic-angle spinning NMR

Rohmer

Lang

Hughes

et al. 2008

Proc. Natl. Acad. Sci. U.S.A.

144

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“…Still, recombinant and tryptic phytochrome fragments, that range either from amino acid residues 1 to 595 (65 kDa) or 65 to 595 (59 kDa) only, i.e. lack the C‐terminal part, display full photoreversibility and kinetics of the P r /P fr photointerconversion similar to the wild‐type photoreceptor [4–6], in accord with the observation that the chromophore structure and the photoinduced structural changes of the recombinant 65‐kDa phytochrome are essentially the same as well [7,8]. However, truncation at position 425 by tryptic digestion substantially affects the formation of P fr , as indicated by the intensity reduction of the long‐wavelength absorption band.…”

Section: Introductionsupporting

confidence: 71%

“…In this work, the 39‐kDa fragment is studied by resonance Raman (RR) spectroscopy that selectively probes the vibrational spectrum of the chromophore [7,9,12–17]. Thus, this technique provides information about the molecular structure of the tetrapyrrole and its interactions with the protein environment.…”

Section: Introductionmentioning

confidence: 99%

Resonance Raman spectroscopic study of the tryptic 39‐kDa fragment of phytochrome

et al. 2000

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The 39-kDa fragment of oat phytochrome phyA, obtained by tryptic digestion at the amino acids 65 and 425, was studied by resonance Raman spectroscopy. The parent state P r reveals far-reaching similarities with that of the native phytochrome implying that the structures of the tetrapyrrole chromophore and its immediate protein environment are not affected by the proteolysis. However, the resonance Raman spectrum of the final product of the P r phototransformation, denoted as P bl , is more closely related to that of the P fr precursor of the native phytochrome, i.e. meta-R C , rather than to that of P fr itself. The resonance Raman spectra indicate a high conformational flexibility of the chromophore in P bl so that, unlike in P fr , the tetrapyrrole rings C and D adopt a largely coplanar conformation. The protein interactions with ring D of the chromophore, which in the native phytochrome stabilize the specific chromophore structure of P fr , cannot be established in the 39-kDa fragment due to the lack of the major C-terminal part of the protein. These findings, furthermore, support the view that the meta-R C C CP fr transition is associated with a coupling of chromophore and protein structural changes that represent crucial events for the photoactivation of phytochrome. ß

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“…When red light is absorbed, PΦB changes its constitution via a Z→E isomerization, which brings the protein into a different conformation, the far‐red light absorbing form (P fr ). The absorption maximum of P fr shows a peak around 730 nm [8]. This red/far‐red light dependent photoconversion is the main characteristic of all phytochrome molecules.…”

Section: Introductionmentioning

confidence: 97%

“…Structural formula of PΦB (A) and PCB (B) [8]. At C‐18 of ring D PΦB has one more double bond (vinyl group) than PCB (ethyl group).…”

Section: Introductionmentioning

confidence: 99%

Arabidopsisphytochromes C and E have different spectral characteristics from those of phytochromes A and B

et al. 2000

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The red/far-red light absorbing phytochromes play a major role as sensor proteins in photomorphogenesis of plants. In Arabidopsis the phytochromes belong to a small gene family of five members, phytochrome A (phyA) to E (phyE). Knowledge of the dynamic properties of the phytochrome molecules is the basis of phytochrome signal transduction research. Beside photoconversion and destruction, dark reversion is a molecular property of some phytochromes. A possible role of dark reversion is the termination of signal transduction. Since Arabidopsis is a model plant for biological and genetic research, we focussed on spectroscopic characterization of Arabidopsis phytochromes, expressed in yeast. For the first time, we were able to determine the relative absorption maxima and minima for a phytochrome C (phyC) as 661/725 nm and for a phyE as 670/724 nm. The spectral characteristics of phyC and E are strictly different from those of phyA and B. Furthermore, we show that both phyC and phyE apoprotein chromophore adducts undergo a strong dark reversion. Difference spectra, monitored with phycocyanobilin and phytochromobilin as the apoprotein's chromophore, and in vivo dark reversion of the Arabidopsis phytochrome apoprotein phycocyanobilin adducts are discussed with respect to their physiological function.z 2000 Federation of European Biochemical Societies.

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Effect of chromophore exchange on the resonance Raman spectra of recombinant phytochromes

Cited by 32 publications

References 19 publications

Light-induced chromophore activity and signal transduction in phytochromes observed by ¹³ C and ¹⁵ N magic-angle spinning NMR

Light-induced chromophore activity and signal transduction in phytochromes observed by ¹³ C and ¹⁵ N magic-angle spinning NMR

Resonance Raman spectroscopic study of the tryptic 39‐kDa fragment of phytochrome

Arabidopsisphytochromes C and E have different spectral characteristics from those of phytochromes A and B

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Effect of chromophore exchange on the resonance Raman spectra of recombinant phytochromes

Cited by 32 publications

References 19 publications

Light-induced chromophore activity and signal transduction in phytochromes observed by 13 C and 15 N magic-angle spinning NMR

Light-induced chromophore activity and signal transduction in phytochromes observed by 13 C and 15 N magic-angle spinning NMR

Resonance Raman spectroscopic study of the tryptic 39‐kDa fragment of phytochrome

Arabidopsisphytochromes C and E have different spectral characteristics from those of phytochromes A and B

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Light-induced chromophore activity and signal transduction in phytochromes observed by ¹³ C and ¹⁵ N magic-angle spinning NMR

Light-induced chromophore activity and signal transduction in phytochromes observed by ¹³ C and ¹⁵ N magic-angle spinning NMR