Locked 5Zs‐biliverdin blocks the Meta‐RA to Meta‐RC transition in the functional cycle of bacteriophytochrome Agp1

Seibeck, Sven; Borucki, Berthold; Otto, H. H.; Inomata, Katsuhiko; Khawn, Htoi; Kinoshita, Hideki; Michael, Norbert; Lamparter, Tilman; Heyn, Maarten P.

doi:10.1016/j.febslet.2007.10.043

Cited by 23 publications

(37 citation statements)

References 21 publications

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“…These changes clearly occur in the subsequent transitions reflecting the Meta-R A to Meta-R C and the Meta-R C to P fr transitions, respectively. In an adduct of Agp1 with a locked 5Zs-BV chromophore the Meta-R A to Meta-R C transition was inhibited (36). The present results provide further evidence for conformational changes of the A-B methine bridge during the Meta-R A to Meta-R C transition.…”

Section: Mechanism Of Probe Attachment and Chromophoresupporting

confidence: 62%

“…The present results do not allow discrimination between these two possibilities. One possible scenario could be that the postulated syn to anti rotation of the C5-C6 single bond (33,36) is associated with a relocation of pyrrole ring A allowing the probe to occupy the initial position of ring A. Alternatively, a partial rotation of ring A could produce a cavity such that the probe slips beneath. These rearrangements could occur for example in the proposed hula-twist motion at C5 that involves a concerted syn to anti rotation and Z to E isomerization (33,58).…”

Section: Mechanism Of Probe Attachment and Chromophorementioning

confidence: 99%

“…Flash photolysis experiments with this adduct suggested that these changes occur in the Meta-R A to Meta-R C transition (36). The stereochemistry of the A-B methine bridge in the P fr state and in the preceding intermediates could not be determined unambiguously yet.…”

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confidence: 99%

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A Polarity Probe for Monitoring Light-induced Structural Changes at the Entrance of the Chromophore Pocket in a Bacterial Phytochrome

Borucki

Lamparter

2009

Journal of Biological Chemistry

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Light-induced structural changes at the entrance of the chromophore pocket of Agp1 phytochrome were investigated by using a thiol-reactive fluorescein derivative that is covalently attached to the genuine chromophore binding site (Cys-20) and serves as a polarity probe. In the apoprotein, the absorption spectrum of bound fluorescein is red-shifted with respect to that of the free label suggesting that the probe enters the hydrophobic chromophore pocket. Assembly of this construct with the chromophores phycocyanobilin or biliverdin is associated with a blue-shift of the fluorescein absorption band indicating the displacement of the probe out of the pocket. The probe does not affect the photochromic and kinetic properties of the noncovalent bilin adducts. Upon photoconversion to P fr , the probe spectrum undergoes again a bathochromic shift and a strong rise in CD indicating a more hydrophobic and asymmetric environment. We propose that the environmental changes of the probe reflect conformational changes at the entrance of the chromophore pocket and are indicative for rearrangements of the chromophore ring A. Flash photolysis measurements showed that the absorption changes of the probe are kinetically coupled to the formation of Meta-R C and P fr . In the biliverdin adduct, an additional component occurs that probably reflects a transition between two Meta-R C substates. Analogous results to that of the noncovalent phycocyanobilin adduct were obtained with the mutant V249C in which probe and chromophore are covalently attached. The conformational changes of the chromophore are correlated to proton transfer to the protein surface.

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Section: Mechanism Of Probe Attachment and Chromophoresupporting

confidence: 62%

Section: Mechanism Of Probe Attachment and Chromophorementioning

confidence: 99%

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A Polarity Probe for Monitoring Light-induced Structural Changes at the Entrance of the Chromophore Pocket in a Bacterial Phytochrome

Borucki

Lamparter

2009

Journal of Biological Chemistry

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“…The chromophore of the Agp1-M15-D197A mutant is only partially protonated at the slightly basic pH used here, whereas in wild type Agp1 the chromophore is completely protonated under these conditions (57). Because the excitation and emission spectra of 15Za-Agp1-M15-D197A are remarkably similar to those of 15Za-Agp1, we propose that fluorescence is related to the protonated chromophore.…”

Section: Discussionmentioning

confidence: 69%

Fluorescence of Phytochrome Adducts with Synthetic Locked Chromophores

Zienicke

Chen

Khawn

et al. 2011

Journal of Biological Chemistry

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We performed steady state fluorescence measurements with phytochromes Agp1 and Agp2 of Agrobacterium tumefaciens and three mutants in which photoconversion is inhibited. These proteins were assembled with the natural chromophore biliverdin (BV), with phycoerythrobilin (PEB), which lacks a double bond in the ring C-D-connecting methine bridge, and with synthetic bilin derivatives in which the ring C-D-connecting methine bridge is locked. All PEB and locked chromophore adducts are photoinactive. According to fluorescence quantum yields, the adducts may be divided into four different groups: wild type BV adducts exhibiting a weak fluorescence, mutant BV adducts with about 10-fold enhanced fluorescence, adducts with locked chromophores in which the fluorescence quantum yields are around 0.02, and PEB adducts with a high quantum yield of around 0.5. Thus, the strong fluorescence of the PEB adducts is not reached by the locked chromophore adducts, although the photoconversion energy dissipation pathway is blocked. We therefore suggest that ring D of the bilin chromophore, which contributes to the extended -electron system of the locked chromophores, provides an energy dissipation pathway that is independent on photoconversion.

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“…For PCB, we have shown that the CD sign of the red transition arises from the facial disposition of the D-ring (22). CD spectra of phytobilin-and BV-containing phytochromes in their P r states share strong negative rotation for the red absorbance band (22)(23)(24)(25). This band exhibits weakly negative rotation in the P fr state of the BphP Agp1 assembled with a synthetic chromophore (25), consistent with structural results suggesting that the D-ring remains ␣-facial during P r /P fr interconversion of biliverdin phytochromes (8,16,18).…”

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confidence: 99%

Distinct classes of red/far-red photochemistry within the phytochrome superfamily

Rockwell

Shang

Martin

et al. 2009

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

134

209

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Phytochromes are a widespread family of photosensory proteins first discovered in plants, which measure the ratio of red to far-red light to control many aspects of growth and development. Phytochromes interconvert between red-absorbing Pr and far-redabsorbing Pfr states via photoisomerization of a covalently-bound linear tetrapyrrole (bilin) chromophore located in a conserved photosensory core. From recent crystal structures of this core region, it has been inferred that the chromophore structures of Pr and Pfr are conserved in most phytochromes. Using circular dichroism spectroscopy and ab initio calculations, we establish that the Pfr states of the biliverdin-containing bacteriophytochromes DrBphP and PaBphP are structurally dissimilar from those of the phytobilincontaining cyanobacterial phytochrome Cph1. This conclusion is further supported by chromophore substitution experiments using semisynthetic bilin monoamides, which indicate that the propionate side chains perform different functional roles in the 2 classes of phytochromes. We propose that different directions of bilin D-ring rotation account for these distinct classes of red/far-red photochemistry.bilin amides ͉ biliprotein ͉ circular dichroism ͉ photoisomerization P hytochromes comprise a class of red/far-red biliprotein photoreceptors that regulate photomorphogenesis, shade avoidance, and development in higher plants (1, 2). Also found in bacteria and fungi, phytochromes characteristically photoconvert between red-absorbing P r and far-red-absorbing P fr states (3,4). Conversion between these 2 states involves Z/E photoisomerization of the 15/16 double bond of their linear tetrapyrrole (bilin) chromophores. Photosensory signaling by phytochromes relies on a conserved core comprising PAS (PER, ARNT, SIM), GAF (cGMP phosphodiesterase, adenylate cyclase, FhlA), and PHY (phytochrome-specific GAF-related) domains (5), with the bilin bound within a conserved pocket of the GAF domain (1, 6). The precise structure of the bilin chromophore, the nature of its covalent linkage to cysteine (Cys) side chains in the protein, and the location of those Cys residues vary among phytochromes (7), and 2 distinct subclasses can be distinguished on these grounds.Phytobilin-containing phytochromes, which include the plant (Phys) and cyanobacterial (Cph1) phytochromes, are found exclusively in oxygenic photosynthetic organisms. The chromophore precursor for these phytochromes is either phytochromobilin or phycocyanobilin (PCB), both of which possess reduced ethylidene-containing A-rings. For these phytochromes, covalent linkage forms between a GAF-domain Cys and the ␣-carbon of the A-ring ethylidene (Fig. S1 A). The much more widespread biliverdin-containing phytochromes (4), which include the bacteriophytochromes (BphPs) and fungal phytochromes, possess covalent linkages between a Cys residue upstream of the PAS domain and the ␤-carbon of the A-ring endo-vinyl group of biliverdin IX␣ (BV; Fig. S1B) (8). P r is commonly the thermally stable dark state for both classes of...

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Locked 5Zs‐biliverdin blocks the Meta‐R_A to Meta‐R_C transition in the functional cycle of bacteriophytochrome Agp1

Cited by 23 publications

References 21 publications

A Polarity Probe for Monitoring Light-induced Structural Changes at the Entrance of the Chromophore Pocket in a Bacterial Phytochrome

A Polarity Probe for Monitoring Light-induced Structural Changes at the Entrance of the Chromophore Pocket in a Bacterial Phytochrome

Fluorescence of Phytochrome Adducts with Synthetic Locked Chromophores

Distinct classes of red/far-red photochemistry within the phytochrome superfamily

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