Absorption Spectra of Photoactive Yellow Protein Chromophores in Vacuum

Nielsen, Ingolf; Boyé-Péronne, Séverine; Abdellahi, Majid; Kristensen, Michael B.; Nielsen, Steen Brøndsted; Andersen, Lars H.

doi:10.1529/biophysj.105.061192

Cited by 98 publications

(157 citation statements)

References 45 publications

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“…As pointed out earlier, double-bond twisting must include the effect of nuclear couplings to other bonds and electronic͞resonance effects. In a very recent report (30), the gas-phase absorption spectrum of the PYP chromophore indicates that the excitation energy is lower than calculated value (22), indicating that an excess vibrational energy is sufficient for double-bond twisting. Moreover, recent molecular dynamics calculations for the protein and in vacuo support the involvement of the double-bond twisting (31).…”

Section: Resultsmentioning

confidence: 85%

Primary steps of the photoactive yellow protein: Isolated chromophore dynamics and protein directed function

Lee

Zewail

2006

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

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The cycle of the photoactive yellow protein (PYP) has been extensively studied, but the dynamics of the isolated chromophore responsible for transduction is unknown. Here, we present realtime observation of the dynamics of the negatively charged chromophore and detection of intermediates along the path of transto-cis isomerization using femtosecond mass selection͞electron detachment techniques. The results show that the role of the protein environment is not in the first step of double-bond twisting (barrier crossing) but in directing efficient conversion to the cisstructure and in impeding radical formation within the protein.femtobiology ͉ transduction ͉ molecular dynamics ͉ photoelectron spectroscopy P hotoactive yellow protein (PYP) is a water-soluble photoreceptor found in Halorhodospira halophila and related halophilic bacterial species (1). The chromophore responsible for perception of light and phototactic response is a deprotonated trans-4-hydroxycinnamic acid (also known as p-coumaric acid) covalently linked to a cysteine residue of the protein via a thioester bond (Fig. 1). Light absorption of the PYP chromophore leads to the initiation of a complex photocycle involving several intermediates. With a variety of biophysical techniques (2-5), it is concluded that the first intermediate (I 0 ) in the photocycle is formed within a few picoseconds, with the chromophore changing to the cis configuration, in a trans-to-cis isomerization process (7-11). The intermediates at longer times have been trapped and identified by x-ray crystallography (12, 13).The critical change of the chromophore in the primary process of isomerization is accompanied or followed by several other processes: proton transfer, protein conformational change, solvation, and disruption of hydrogen-bonding. This complexity can be reduced if the chromophore in its true anionic, not neutral, structure can be studied free of perturbations. With model compounds, the solvent effect can be assessed by studying them in the solution phase (14-18). However, the nature of intramolecular change and the timescales involved are still unknown. It is, therefore, desirable to study the primary isomerization dynamics of the PYP chromophore in isolation, especially in view of the fact that the mechanism for forming the first intermediate (I 0 ) is debatable (2). A gas-phase spectroscopic study has already been reported (19), but it was for the neutral molecule, not the anion structure in the protein.In this work, we report direct observation of the dynamics of the isolated PYP anionic chromophore. To disentangle the role of protein binding and distant torsions, we preserved the central structure involved in the isomerization about the double bond but use the CH 3 group for termination (see Fig. 1). The chromophore, hereafter called P, is excited with a femtosecond pulse from its ground state, the dark state in the protein, to the trans configuration using the mass-selected ions. For probing we use another femtosecond pulse to photodetach and resolve the phot...

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Section: Resultsmentioning

confidence: 85%

Primary steps of the photoactive yellow protein: Isolated chromophore dynamics and protein directed function

Lee

Zewail

2006

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

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“…[14] The gas-phase absorption spectra were recorded at the electrostatic ion storage ring ELISA, [15] which has previously been used to study photoabsorption of biological chromophores (see, for example, reference [16]), and in particular those of the PYP. [9,10,17] Briefly, deprotonated ions of the three PYP chromophore derivatives (shown in Scheme 1) were produced in the gas phase by using electrospray ionization. After pretrapping for up to 100 ms in a multipole ion trap with room temperature and having He as the buffer gas, a bunch of ions were accelerated to 22 keV, magnetically mass selected, and injected into ELISA.…”

Section: Experimental and Computational Methodsmentioning

confidence: 99%

“…[1] In the case of PYP, the absorption of pCA À in aqueous phase is strongly blueshifted with respect to the gas-phase spectrum: it presents a maximum of absorption at 4.35 eV (285 nm) and a shoulder at 3.97 eV (312 nm). [9] The absence of perturbations from a surrounding medium makes gas-phase absorption spectra a better reference than those obtained in solution for evaluating protein tuning and, importantly, for tests of ab initio calculations.…”

Section: Introductionmentioning

confidence: 99%

Spectroscopic Implications of the Electron Donor–Acceptor Effect in the Photoactive Yellow Protein Chromophore

Rocha‐Rinza

Christiansen

Rahbek

et al. 2010

Chemistry A European J

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The importance of the donor-acceptor push-pull system in the photoabsorption of the trans p-coumaric acid, the cofactor within the photoactive yellow protein and other xanthopsins, has been investigated. We recorded gas-phase absorption spectra and performed high-level quantum chemical calculations of three chromophore models, namely, the deprotonated trans ortho-, meta- and para-methyl coumarates. The ortho and para isomers, which have the electron-donating phenoxy oxygen and the electron-withdrawing acyl group in conjugation, present absorptions in the high-energy region of the visible spectrum, that is, in the interval of wavelengths in which the photoactivity of the xanthopsins is observed. On the other hand, the meta isomer, in which the conjugation between the phenoxy and acyl groups is disrupted, exhibits a significantly shifted maximum and presents no absorption in the region from blue to ultraviolet A. It is found that the push-pull system in the trans p-coumaric acid is critical for the wavelength and the intensity of its photoabsorption. Absorption spectra were also measured in methanol and showed an appreciable hypsochromic effect. Linear response calculations within the formalism of the approximate coupled cluster singles and doubles CC2 model and time-dependent DFT using the functional CAM-B3LYP provided insights into the relevant processes of excitation and aided to the interpretation of the experimental results. There is good agreement between theory and experiment in the description of the gas-phase absorption spectra of the considered chromophore models. Differential density plots were used to predict the effect of hydrogen-bonded amino acids to the trans p-coumaric acid on the protein tuning of this chromophore.

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“…[35] Steady-state absorption spectra of anionic p-coumaric acid and a thioester analogue have been recently measured in vacuo. [39] A time-resolved study of another isolated analogue was recently reported, showing that the initial twisting motion occurs on the same timescale in the gas-phase as inside the protein, though subsequent channeling through the conical intersection is less efficient, owing to the need for excess energy redistribution by intramolecular vibrational relaxation (IVR). [40] p-Coumaric acid is present in its anionic form in PYP.…”

Section: Introductionmentioning

confidence: 99%

Ultrafast Photoisomerization of Photoactive Yellow Protein Chromophore Analogues in Solution: Influence of the Protonation State

et al. 2006

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We investigate solvent viscosity and polarity effects on the photoisomerization of the protonated and deprotonated forms of two analogues of the photoactive yellow protein (PYP) chromophore. These are trans-p-hydroxybenzylidene acetone and trans-p-hydroxyphenyl cinnamate, studied in solutions of different polarity and viscosity at room temperature, by means of femtosecond fluorescence up-conversion. The fluorescence lifetimes of the protonated forms are found to be barely sensitive to solvent viscosity, and to increase with increasing solvent polarity. In contrast, the fluorescence decays of the deprotonated forms are significantly slowed down in viscous media and accelerated in polar solvents. These results elucidate the dramatic influence of the protonation state of the PYP chromophore analogues on their photoinduced dynamics. The viscosity and polarity effects are, respectively, interpreted in terms of different isomerization coordinates and charge redistribution in S(1). A trans-to-cis isomerization mechanism involving mainly the ethylenic double-bond torsion and/or solvation is proposed for the anionic forms, whereas "concerted" intramolecular motions are proposed for the neutral forms.

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Absorption Spectra of Photoactive Yellow Protein Chromophores in Vacuum

Cited by 98 publications

References 45 publications

Primary steps of the photoactive yellow protein: Isolated chromophore dynamics and protein directed function

Primary steps of the photoactive yellow protein: Isolated chromophore dynamics and protein directed function

Spectroscopic Implications of the Electron Donor–Acceptor Effect in the Photoactive Yellow Protein Chromophore

Ultrafast Photoisomerization of Photoactive Yellow Protein Chromophore Analogues in Solution: Influence of the Protonation State

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