When Is the Helix Conformation Restored after the Reverse Reaction of Phototropin?

Kawaguchi, Yuki; Nakasone, Yusuke; Tokutomi, Satoru; Terazima, Masahide

doi:10.1021/ja1020519

Cited by 8 publications

(6 citation statements)

References 21 publications

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“…This may be due to probe light‐induced acceleration of thermal recovery in these proteins as the 1.8‐ANS fluorescence has to be excited at relatively short wavelength ( i.e. 350 nm; compare [26,27]).…”

Section: Resultsmentioning

confidence: 99%

On the Binding of BODIPY‐GTP by the Photosensory Protein YtvA from the Common Soil Bacterium Bacillus subtilis^†

Nakasone

Hellingwerf²

2011

Photochem & Photobiology

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The YtvA protein, which is one of the proteins that comprises the network carrying out the signal transfer inducing the general stress response in Bacillus subtilis, is composed of an N-terminal LOV domain (that binds a flavin [FMN]) and a C-terminal STAS domain. This latter domain shows sequence features typical for a nucleotide (NTP) binding protein. It has been proposed (FEBS Lett., 580 [2006], 3818) that BODIPY-GTP can be used as a reporter for nucleotide binding to this site and that activation of the LOV domain by blue light is reflected in an alteration of the BODIPY-GTP fluorescence. Here we confirm that BODIPY-GTP indeed binds to YtvA, but rather nonspecifically, and not limited to the STAS domain. Blue-light modulation of fluorescence emission of YtvA-bound BODIPY-GTP is observed both in the full-length YtvA protein and in a truncated protein composed of the LOV-domain plus the LOV-STAS linker region (YtvA(1-147)) as a light-induced decrease in fluorescence emission. The isolated LOV domain (i.e. without the linker region) does not show such BODIPY-GTP fluorescence changes. Dialysis experiments have confirmed the blue-light-induced release of BODIPY-GTP from YtvA.

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Section: Resultsmentioning

confidence: 99%

On the Binding of BODIPY‐GTP by the Photosensory Protein YtvA from the Common Soil Bacterium Bacillus subtilis^†

Nakasone

Hellingwerf²

2011

Photochem & Photobiology

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“…Time-resolved nuclear magnetic resonance (NMR) spectroscopy on the isolated LOV2 domain from Avena sativa phototropin1 has revealed a difference between the mean time constant resulting from NMR studies and the lifetime of the adduct state by a factor of 1.2, but in the time region of seconds 38 . Moreover, the recovery of the LOV2-linker from Arabidopsis takes place rapidly with a 13 ms time constant according to transient grating experiments 39 . A difference on the time scale of minutes has only been reported previously for a LOV domain with flanking helices, PpSB1-LOV-R66I from Pseudomonas putida , by time-course SAXS experiments 22 …”

Section: Discussionmentioning

confidence: 99%

Arguments for an additional long-lived intermediate in the photocycle of the full-length aureochrome 1c receptor: A time-resolved small-angle X-ray scattering study

Bannister

Böhm

Zinn

et al. 2019

Structural Dynamics

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Aureochromes (AUREO) act as blue-light photoreceptors in algae. They consist of a light-, oxygen-, voltage-sensitive (LOV) domain and a DNA-binding basic region/leucine zipper. Illumination of the flavin cofactor in LOV leads to the formation of an adduct, followed by global structural changes. Here, we first applied UV/vis spectroscopy to characterize the photocycle of full-length aureochrome 1c ( Pt AUREO1c) from the diatom Phaeodactylum tricornutum . With a time constant of 850 s and a quantum yield of 23%, Pt AUREO1c reveals a faster recovery time and a much lower sensitivity toward light than Pt AUREO1a, pointing to its role as a high light sensor in vivo . UV/vis spectroscopy offers details on the local recovery of the flavin chromophore. However, kinetic information on the global structural recovery of full-length AUREO or any other multidomain LOV protein is missing. This information is essential not least for the photoreceptors' applications as optogenetic devices. Therefore, we established a procedure to apply small-angle X-ray scattering on Pt AUREO1c in a time-resolved manner employing an in-house setup. In combination with UV/vis spectroscopy under similar conditions, we revealed a discrepancy between the recovery of the global protein structure and the adduct lifetime. Accordingly, we propose to supplement the photocycle by an intermediate state (I447), which decays with a time constant of about 800 s and prolongs the lifetime of the signaling state.

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“…Thus, absorption of blue light leads to the transient formation of a covalent bond between the FMN cofactor and the protein, which slowly ruptures in the dark to regenerate the noncovalent dark ground state species, presumably through a base‐catalyzed mechanism (33,34). Absorption of near‐UV light may also rupture the covalent adduct (35,36). Formation of a covalent bond to FMN bond triggers protein conformational changes on the surface of the PAS core which weaken interactions of this core with a C‐terminal amphiphilic helix called Jα, packed against its central β‐sheet (37–39).…”

Section: Introductionmentioning

confidence: 99%

The Primary Photophysics of the Avena sativa Phototropin 1 LOV2 Domain Observed with Time‐resolved Emission Spectroscopy^†

Stokkum

Gauden

Crosson

et al. 2011

Photochem & Photobiology

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The phototropins are blue-light receptors that base their light-dependent action on the reversible formation of a covalent bond between a flavin mononucleotide (FMN) cofactor and a conserved cysteine in light, oxygen or voltage (LOV) domains. The primary reactions of the Avena sativa phototropin 1 LOV2 domain were investigated by means of time-resolved and low-temperature fluorescence spectroscopy. Synchroscan streak camera experiments revealed a fluorescence lifetime of 2.2 ns in LOV2. A weak long-lived component with emission intensity from 600 to 650 nm was assigned to phosphorescence from the reactive FMN triplet state. This observation allowed determination of the LOV2 triplet state energy level at physiological temperature at 16600 cm−1. FMN dissolved in aqueous solution showed pH-dependent fluorescence lifetimes of 2.7 ns at pH 2 and 3.9–4.1 ns at pH 3 to 8. Here, too, a weak phosphorescence band was observed. The fluorescence quantum yield of LOV2 increased from 0.13 to 0.41 upon cooling the sample from 293 to 77 K. A pronounced phosphorescence emission around 600 nm was observed in the LOV2 domain between 77 and 120 K in the steady-state emission.

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When Is the Helix Conformation Restored after the Reverse Reaction of Phototropin?

Cited by 8 publications

References 21 publications

On the Binding of BODIPY‐GTP by the Photosensory Protein YtvA from the Common Soil Bacterium Bacillus subtilis^†

On the Binding of BODIPY‐GTP by the Photosensory Protein YtvA from the Common Soil Bacterium Bacillus subtilis^†

Arguments for an additional long-lived intermediate in the photocycle of the full-length aureochrome 1c receptor: A time-resolved small-angle X-ray scattering study

The Primary Photophysics of the Avena sativa Phototropin 1 LOV2 Domain Observed with Time‐resolved Emission Spectroscopy^†

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When Is the Helix Conformation Restored after the Reverse Reaction of Phototropin?

Cited by 8 publications

References 21 publications

On the Binding of BODIPY‐GTP by the Photosensory Protein YtvA from the Common Soil Bacterium Bacillus subtilis†

On the Binding of BODIPY‐GTP by the Photosensory Protein YtvA from the Common Soil Bacterium Bacillus subtilis†

Arguments for an additional long-lived intermediate in the photocycle of the full-length aureochrome 1c receptor: A time-resolved small-angle X-ray scattering study

The Primary Photophysics of the Avena sativa Phototropin 1 LOV2 Domain Observed with Time‐resolved Emission Spectroscopy†

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Product

Resources

About

On the Binding of BODIPY‐GTP by the Photosensory Protein YtvA from the Common Soil Bacterium Bacillus subtilis^†

On the Binding of BODIPY‐GTP by the Photosensory Protein YtvA from the Common Soil Bacterium Bacillus subtilis^†

The Primary Photophysics of the Avena sativa Phototropin 1 LOV2 Domain Observed with Time‐resolved Emission Spectroscopy^†