The Phytochromes

Tu, Shih-Long; Lagarias, J. Clark

doi:10.1002/352760510x.ch6

Cited by 26 publications

(29 citation statements)

References 126 publications

(187 reference statements)

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“…This conversion is predicted to involve a Z to E isomerization of the C15-C16 methine double bond between the C and D pyrrole rings of the chromophore (16 -18). On the microsecond time scale, thermal relaxation steps lead to the formation of the Meta-R a intermediate with a maximum absorption at 663 nm (19). The Meta-R a intermediate in turn decays in micro-to milliseconds to a deprotonated Meta-R c intermediate with an absorption maximum at 725 nm (20,21).…”

mentioning

confidence: 99%

Mutational Analysis of Deinococcus radiodurans Bacteriophytochrome Reveals Key Amino Acids Necessary for the Photochromicity and Proton Exchange Cycle of Phytochromes

Wagner

Zhang

Stetten

et al. 2008

Journal of Biological Chemistry

196

353

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The ability of phytochromes (Phy) to act as photointerconvertible light switches in plants and microorganisms depends on key interactions between the bilin chromophore and the apoprotein that promote bilin attachment and photointerconversion between the spectrally distinct red light-absorbing Pr conformer and far red light-absorbing Pfr conformer. Using structurally guided site-directed mutagenesis combined with several spectroscopic methods, we examined the roles of conserved amino acids within the bilin-binding domain of Deinococcus radiodurans bacteriophytochrome with respect to chromophore ligation and Pr/Pfr photoconversion. Incorporation of biliverdin IX␣ (BV), its structure in the Pr state, and its ability to photoisomerize to the first photocycle intermediate are insensitive to most single mutations, implying that these properties are robust with respect to small structural/electrostatic alterations in the binding pocket. In contrast, photoconversion to Pfr is highly sensitive to the chromophore environment. Many of the variants form spectrally bleached Meta-type intermediates in red light that do not relax to Pfr. Particularly important are Asp-207 and His-260, which are invariant within the Phy superfamily and participate in a unique hydrogen bond matrix involving the A, B, and C pyrrole ring nitrogens of BV and their associated pyrrole water. Resonance Raman spectroscopy demonstrates that substitutions of these residues disrupt the Pr to Pfr protonation cycle of BV with the chromophore locked in a deprotonated Meta-R c -like photoconversion intermediate after red light irradiation. Collectively, the data show that a number of contacts contribute to the unique photochromicity of Phy-type photoreceptors. These include residues that fix the bilin in the pocket, coordinate the pyrrole water, and possibly promote the proton exchange cycle during photoconversion.The phytochrome (Phy) 5 superfamily encompasses a large and diverse set of photoreceptors present in the plant, fungal, and bacterial kingdoms where they play critical roles in various light-regulated processes (1-3). These processes range from the control of phototaxis, pigmentation, and photosynthetic potential in proteobacteria and cyanobacteria to seed germination, chloroplast development, shade avoidance, and flowering time in higher plants. Phys are unique among photoreceptors in being able to assume two stable, photointerconvertible conformers, designated Pr and Pfr based on their respective absorption maxima in the red and far-red spectral regions. By cycling between Pr and Pfr, Phys act as light-regulated switches in various photosensory cascades.Phys are homodimeric complexes with each polypeptide containing a single bilin (or linear tetrapyrrole) chromophore, which binds autocatalytically via a thioether linkage to a positionally conserved cysteine (1-3). The photosensing portion typically contains Per/Arndt/Sim (PAS) and cGMP phosphodiesterase/adenyl cyclase/FhlA (GAF) domains, which are essential for bilin binding and Pr assembly, and t...

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mentioning

confidence: 99%

Mutational Analysis of Deinococcus radiodurans Bacteriophytochrome Reveals Key Amino Acids Necessary for the Photochromicity and Proton Exchange Cycle of Phytochromes

Wagner

Zhang

Stetten

et al. 2008

Journal of Biological Chemistry

196

353

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“…These two forms are denoted according to the absorption maxima as red-absorbing form Pr and far-red absorbing form Pfr. Upon light absorption in the Pr state, the tetrapyrrole chromophore undergoes a double bond Z 3 E isomerization at the C-D methine bridge to yield the photoproduct lumi-R, which thermally relaxes via several meta intermediate states to the Pfr form (3).…”

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confidence: 99%

Highly Conserved Residues Asp-197 and His-250 in Agp1 Phytochrome Control the Proton Affinity of the Chromophore and Pfr Formation

Stetten

Seibeck

Michael

et al. 2007

Journal of Biological Chemistry

111

214

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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 3 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.

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“…phys exist in two interconvertible forms: the inactive Pr and the biologically active Pfr form. Absorption of red-light (R) photons by the photoreceptor, a polypeptide with a tetrapyrrole chromophore, induces a conformational change from the Pr to Pfr form (activation); absorption of far-red light (FR) by the Pfr form triggers the converse process (2). After photoactivation, phys move from the cytosol to the nucleus and induce rapid changes in target gene expression (3).…”

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confidence: 99%

Mechanistic duality of transcription factor function in phytochrome signaling

Al-Sady

Kikis

Monte

et al. 2008

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

102

147

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The phytochrome (phy) family of sensory photoreceptors (phyA-E in Arabidopsis) elicit changes in gene expression after lightinduced migration to the nucleus, where they interact with basic helix-loop-helix transcription factors, such as phytochrome-interacting factor 3 (PIF3). The mechanism by which PIF3 relays phy signals, both early after initial light exposure and later during long-term irradiation, is not understood. Using transgenically expressed PIF3 variants, carrying site-specific amino acid substitutions that block the protein from binding either to DNA, phyA, and/or phyB, we examined the involvement of PIF3 in early, phy-induced marker gene expression and in modulating long-term, phy-imposed inhibition of hypocotyl cell elongation under prolonged, continuous irradiation. We describe an unanticipated dual mechanism of PIF3 action that involves the temporal uncoupling of its two most central molecular functions. We find that in early signaling, PIF3 acts positively as a transcription factor, exclusively requiring its DNA-binding capacity. Contrary to previous proposals, PIF3 functions as a constitutive coactivator in this process, without the need for phy binding and subsequent phy-induced modifications. This finding implies that another factor(s) is conditionally activated by phy and functions in concert with PIF3, to induce target gene transcription. In contrast, during long-term irradiations, PIF3 acts exclusively through its phyB-interacting capacity to control hypocotyl cell elongation, independently of its ability to bind DNA. Unexpectedly, PIF3 uses this capacity to regulate phyB protein abundance (and thereby global photosensory sensitivity) to modulate this long-term response rather than participating directly in the transduction chain as a signaling intermediate.phytochrome-interacting factor 3 ͉ photosensory receptor ͉ constitutive coactivator ͉ posttranslational regulation ͉ ELIP2

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The Phytochromes

Cited by 26 publications

References 126 publications

Mutational Analysis of Deinococcus radiodurans Bacteriophytochrome Reveals Key Amino Acids Necessary for the Photochromicity and Proton Exchange Cycle of Phytochromes

Mutational Analysis of Deinococcus radiodurans Bacteriophytochrome Reveals Key Amino Acids Necessary for the Photochromicity and Proton Exchange Cycle of Phytochromes

Highly Conserved Residues Asp-197 and His-250 in Agp1 Phytochrome Control the Proton Affinity of the Chromophore and Pfr Formation

Mechanistic duality of transcription factor function in phytochrome signaling

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