Light-induced Structural Changes Research Articles

Large Deformation of Helix F during the Photoreaction Cycle of Pharaonis Halorhodopsin in Complex with Azide

Halorhodopsin from Natronomonas pharaonis (pHR), a retinylidene protein that functions as a light-driven chloride ion pump, is converted into a proton pump in the presence of azide ion. To clarify this conversion, we investigated light-induced structural changes in pHR using a C2 crystal that was prepared in the presence of Cl− and subsequently soaked in a solution containing azide ion. When the pHR-azide complex was illuminated at pH 9, a profound outward movement (∼4 Å) of the cytoplasmic half of helix F was observed in a subunit with the EF loop facing an open space. This movement created a long water channel between the retinal Schiff base and the cytoplasmic surface, along which a proton could be transported. Meanwhile, the middle moiety of helix C moved inward, leading to shrinkage of the primary anion-binding site (site I), and the azide molecule in site I was expelled out to the extracellular medium. The results suggest that the cytoplasmic half of helix F and the middle moiety of helix C act as different types of valves for active proton transport.

2SAP-02 ロドプシン群蛋白質の光誘起構造変化に関するX線結晶解析(2SAP ロドプシン研究の過去・現在・未来,シンポジウム,日本生物物理学会第51回年会(2013年度))

2SAP-02 ロドプシン群蛋白質の光誘起構造変化に関するX線結晶解析(2SAP ロドプシン研究の過去・現在・未来,シンポジウム,日本生物物理学会第51回年会(2013年度))

Light-induced reactivation of O2-tolerant membrane-bound [Ni–Fe] hydrogenase from the hyperthermophilic bacterium Aquifex aeolicus under turnover conditions

We report the effect of UV-Vis light on the membrane-bound [Ni-Fe] hydrogenase from Aquifex aeolicus under turnover conditions. Using electrochemistry, we show a potential dependent light sensitivity and propose that a light-induced structural change of the [Ni-Fe] active site is related to an enhanced reactivation of the hydrogenase under illumination at high potentials.

Comparative FTIR Study of a New Fungal Rhodopsin

Bacteriorhodopsin (BR) is a light-driven proton pump of halophilic Archaea , and BR-like proton-pumping rhodopsins have been discovered in Bacteria and Eucarya as well. Leptosphaeria rhodopsin (LR) and Phaeosphaeria Rhodopsin 2 (PhaeoRD2) are both fungal rhodopsins in such a functional class, even though they belong to different branches of the phylogenetic tree. In this study, we compared light-induced structural changes in the K, L, and M photointermediates for PhaeoRD2, LR, and BR using low-temperature Fourier transform infrared (FTIR) spectroscopy. We observed a strongly hydrogen-bonded water molecule in PhaeoRD2 (water O-D stretch in D(2)O at 2258 cm(-1)) as well as in LR and BR. This observation provided additional experimental evidence to the concept that strongly hydrogen-bonded water molecule is the functional determinant of light-driven proton pumping. The difference FTIR spectra for all the K, L, and M states are surprisingly similar between PhaeoRD2 and LR, but not for BR. PhaeoRD2 is more homologous to LR than to BR, but the difference is small. The amino acid identities between PhaeoRD2 and LR, and between PhaeoRD2 and BR are 34.5% and 30.2%, respectively. In addition, the amino acids uniquely identical for the fungal rhodopsins are located rather far from the retinal chromophore. In fact, the amino acid identities within 4 Å from retinal are the same among PhaeoRD2, LR, and BR. For more than 100 amino acids located within 12 Å from retinal, the identities are 48.7% between PhaeoRD2 and LR, 46.0% between PhaeoRD2 and BR, and 47.8% between LR and BR. These results suggest that protein core structures are equally different among the three rhodopsins. Thus, the identical FTIR spectra between PhaeoRD2 and LR (but not BR), even for the K state, indicate that fungal rhodopsins possess some common structural motif and dynamics not obvious from the amino acid sequences.

Small-angle neutron scattering study of the ultrastructure of chloroplast thylakoid membranes — Periodicity and structural flexibility of the stroma lamellae

The multilamellar organization of freshly isolated spinach and pea chloroplast thylakoid membranes was studied using small-angle neutron scattering. A broad peak at ~0.02Å−1 is ascribed to diffraction from domains of ordered, unappressed stroma lamellae, revealing a repeat distance of 294ű7Å in spinach and 345ű11Å in pea. The peak position and hence the repeat distance of stroma lamellae is strongly dependent on the osmolarity and the ionic strength of the suspension medium, as demonstrated by varying the sorbitol and the Mg++-concentration in the sample. For pea thylakoid membranes, we show that the repeat distance decreases when illuminating the sample with white light, in accordance with our earlier results on spinach, also regarding the observation that addition of an uncoupler prohibits the light-induced structural changes, a strong indication that these changes are driven by the transmembrane proton gradient. We show that the magnitude of the shrinkage is strongly dependent on light intensity and that the repeat distance characteristic of the dark state after illumination is different from the initial dark state. Prolonged strong illumination leads to irreversible changes and swelling as reflected in increased repeat distances. The observed reorganizations are discussed within the frames of the current structural models of the granum-stroma thylakoid membrane assembly and the regulatory mechanisms in response to variations in the environmental conditions in vivo. This article is part of a Special Issue entitled: Photosynthesis Research for Sustainability: from Natural to Artificial.

Lipid Binding to the Carotenoid Binding Site in Photosynthetic Reaction Centers

Lipid binding to the carotenoid binding site near the inactive bacteriochlorophyll monomer was probed in the reaction centers of carotenoid-less mutant, R-26 from Rhodobacter sphaeroides. Recently, a marked light-induced change of the local dielectric constant in the vicinity of the inactive bacteriochlorophyll monomer was reported in wild type that was attributed to structural changes that ultimately lengthened the lifetime of the charge-separated state by 3 orders of magnitude (Deshmukh, S. S.; Williams, J. C.; Allen, J. P.; Kalman, L. Biochemistry 2011, 50, 340). Here in the R-26 reaction centers, the combination of light-induced structural changes and lipid binding resulted in a 5 orders of magnitude increase in the lifetime of the charge-separated state involving the oxidized dimer and the reduced primary quinone in proteoliposomes. Only saturated phospholipids with fatty acid chains of 12 and 14 carbon atoms long were bound successfully at 8 °C by cooling the reaction center protein slowly from room temperature. In addition to reporting a dramatic increase of the lifetime of the charge-separated state at physiologically relevant temperatures, this study reveals a novel lipid binding site in photosynthetic reaction center. These results shed light on a new potential application of the reaction center in energy storage as a light-driven biocapacitor since the charges separated by ∼30 Å in a low-dielectric medium can be prevented from recombination for hours.

Light-Induced Conformational Changes in Photosynthetic Reaction Centers: Impact of Detergents and Lipids on the Electronic Structure of the Primary Electron Donor

Light-induced hypsochromic shifts of the Q(y) absorption band of the bacteriochlorophyll dimer (P) from 865 to 850 nm were identified using continuous illumination of dark-adapted reaction centers (RCs) from Rhodobacter capsulatus when dispersed in the most commonly used detergent, the zwitterionic lauryl N-dimethylamine-N-oxide. Such a shift is known to be the consequence of the decreased degree of delocalization of P. A 2-fold acceleration of the recovery kinetics of P(+) was found in RCs that underwent light-induced structural changes compared to those where the P-band position did not change. The light-induced shift was irreversible except in the presence of a secondary electron donor. Prolonged (15 min) illumination resulted in a shift in the position of the P-band even in neutral or negatively charged detergents. In contrast, RCs reconstituted into liposomes made from lipids with different headgroup charges showed light-induced shifts only if shorter fatty acid chains were used. The light-induced conformational changes caused a prominent decrease of the redox potential of P ranging from 120 to 160 mV depending on the detergent compared to the potential of P in dark-adapted reaction centers. The measured light-induced potential decreases were 55 to 85 mV larger than those reported for reaction centers where the P-band position remained at 865 nm. The influence of structural factors, such as the delocalization of the electron hole on P(+), the involvement of Tyr M210, and the hydrophobic mismatch between the thickness of the hydrophobic belt of the detergent micelles or the lipid bilayer and the RC protein, on the spectral features and electron transfer kinetics is discussed.

Light-induced structural changes in Eu-doped (Pb,La)(Zr,Ti)O3 ceramics

Fluorescence spectra of poled and unpoled Eu-doped (Pb,La)(Zr,Ti)O3 ceramics with various compositions were measured using 580 nm excitation. The ratio of the intensities of the Eu3+ D50→F72/D50→F71 fluorescence transitions was used as an indicator for the symmetry of the Eu3+ site. As the symmetry decreases, the ratio increases. Increasing the Zr content in (Pb,La)(Zr,Ti)O3 increases the ratio, indicating that the symmetry of the Eu3+ site decreases. Exposure of the Eu-doped (Pb,La)(Zr,Ti)O3 samples to UV light also increases the intensity ratio, indicating that UV light exposure leads to structural distortions in Eu-doped (Pb,La)(Zr,Ti)O3 samples, lowering the Eu3+ site symmetry.

Light-Induced Conformational Changes in Photosynthetic Reaction Centers: Redox-Regulated Proton Pathway near the Dimer

The influence of the hydrogen bonds on the light-induced structural changes were studied in the wild type and 11 mutants with different hydrogen bonding patterns of the primary electron donor of reaction centers from Rhodobacter sphaeroides. Previously, using the same set of mutants at pH 8, a marked light-induced change of the local dielectric constant in the vicinity of the dimer was reported in wild type and in mutants retaining Leu L131 that correlated with the recovery kinetics of the charge-separated state [ Deshmukh et al. (2011) Biochemistry, 50, 340-348]. In this work after prolonged illumination the recovery of the oxidized dimer was found to be multiphasic in all mutants. The fraction of the slowest phase, assigned to a recovery from a conformationally altered state, was strongly pH dependent and found to be extremely long at room temperature, at pH 6, with rate constants of ∼10(-3) s(-1). In wild type and in mutants with Leu at L131 the very long recovery kinetics was coupled to a large proton release at pH 6 and a decrease of up to 79 mV of the oxidation potential of the dimer. In contrast, in the mutants carrying the Leu to His mutation at the L131 position, only a negligible fraction of the dimer exhibited lowered potential, the large proton release was not observed, the oxidized dimer recovered 1 or 2 orders of magnitude faster depending on the pH, and the very long-lived state was not or barely detectable. These results are modeled as arising from the loss of a proton pathway from the bacteriochlorophyll dimer to the solvent when His is present at the L131 position.

Electron−Nuclear and Electron−Electron Double Resonance Spectroscopies Show that the Primary Quinone Acceptor QA in Reaction Centers from Photosynthetic Bacteria Rhodobacter sphaeroides Remains in the Same Orientation Upon Light-Induced Reduction

Reaction centers (RCs) from the photosynthetic bacterium Rhodobacter (Rb.) sphaeroides R-26 exhibit changes in the recombination kinetics of the charge-separated radical-pair state, P(·+) Q(A)(·-), composed of the dimeric bacteriochlorophyll donor P and the ubiquinone-10 acceptor Q(A), depending on whether the RCs are cooled to cryogenic temperatures in the dark or under continuous illumination (Kleinfeld et al. Biochemistry 1984, 23, 5780-5786). Structural changes near redox-active cofactors have been postulated to be responsible for these changes in kinetics and to occur in the course of light-induced oxidation and reduction of the cofactors thereby assuring a high quantum yield. Here we investigated such potential light-induced structural changes, associated with the formation of P(·+) Q(A)(·-), via pulsed electron-nuclear double resonance (ENDOR) at Q-band (34 GHz) and pulsed electron-electron double resonance (PELDOR) at W-band (95 GHz). Two types of light excitation have been employed for which identical RC samples were prepared: (a) one sample was frozen in the dark and then illuminated to generate transient P(·+) Q(A)(·-), and (b) one was frozen under illumination which resulted in both trapped and transient P(·+) Q(A)(·-) at 80 K. The hyperfine interactions between Q(A)(·-) and the protein were found to be the same in RCs frozen in the dark as in RCs frozen under illumination. Furthermore, these interactions are completely consistent with those observed in RC crystals frozen in the dark. Thus, QA remains in its binding site with the same position and orientation upon reduction. This conclusion is consistent with the result of our orientation-resolving PELDOR experiments on transient P(·+) Q(A)(·-) radical pairs. However, these findings are incompatible with the recently proposed ~60° reorientation of Q(A) upon its photoreduction, as deduced from an analysis of Q-band quantum-beat oscillations (Heinen et al. J. Am. Chem. Soc. 2007, 129, 15935-15946). Such a large reorientation appears improbable, and our objections against this proposition are substantiated here in detail. Our results show that Q(A) is initially in an orientation that is favorable for its light-driven reduction. This diminishes the reorganization requirements for fast electron reduction and high quantum efficiency.

Isolation and characterization of lamellar aggregates of LHCII and LHCII-lipid macro-assemblies with light-inducible structural transitions.

We describe the method of isolation of loosely stacked lamellar aggregates of LHCII that are capable of undergoing light-induced reversible structural changes, similar to those in granal thylakoid membranes (LHCII, the main chlorophyll a/b light-harvesting antenna complex of photosystem II). This unexpected structural flexibility of the antenna complexes depends largely on the lipid content that is retained during the isolation. As revealed by circular dichroism, in lipid-LHCII aggregates, the pigment-pigment interactions are very similar to those in the thylakoid membranes, while they differ significantly from those in solubilized trimers. The essence of the procedure is to adjust--for the plant material used--the proper conditions of detergent solubilization and purification that are mild enough for the associated lipids but provide sufficient purity. Microcrystals and most other LHCII preparations, which are more delipidated, are not capable of similar changes. The light-induced structural reorganizations can be enhanced by the addition of different thylakoid lipids, which--depending on the lipid species--also lead to the transformation of the lamellar structure. The preparation of different LHCII-lipid macro-assemblies is also described. Both in structurally flexible LHCII preparations and in thylakoids, the changes originate from a thermo-optic effect: fast local thermal transients, T-jumps, due to the dissipation of the (excess) excitation energy, which lead to elementary structural transitions in the close vicinity of the dissipating centers. This can occur because thylakoids and structurally flexible LHCII assemblies, but, e.g., not the microcrystals, exhibit a thermal instability below the denaturation temperature, and thus (local) heating leads to reorganizations without the loss of the molecular architecture of the constituents. We also list the main biochemical and biophysical techniques that can be used for testing the structural flexibility of LHCII, and discuss the potential physiological significance of the structural changes in light adaptation and photoprotection of plants.

Coupling between the BLUF and EAL domains in the blue light-regulated phosphodiesterase BlrP1

The first biochemical and structural characterization of the full-length active photoreceptor BlrP1 from Klebsiella pneumoniae was recently reported by Barends et al. [Nature 459:1015-1018, (2009)]. The light-regulated catalytic function of its C-terminal c-di-guanosine monophosphate phosphodiesterase, the EAL (Glu-Ala-Leu) domain, is activated by the N-terminal sensor of blue light using the flavin adenine dinucleotide (BLUF) domain. We performed molecular dynamics simulations on the dimeric BlrP1 protein in order to examine the coupling regions that are presumably involved in transmitting light-induced structural changes which occur in the BLUF domain to the EAL domain. According to the results of simulations and an analysis of the hydrogen bonding between the respective polypeptide chains, the region containing the site on the α3α4 loop of BLUF is responsible for communication between the photosensing and catalytic domains in the dimeric BlrP1 protein.

Light-induced structural changes in photosynthetic reaction centres

Light-induced structural changes in photosynthetic reaction centres

Quantum Chemistry Calculations Provide Support to the Mechanism of the Light-Induced Structural Changes in the Flavin-Binding Photoreceptor Proteins.

The proposed mechanisms of photoinduced reactions in the blue light using flavin chromophore photoreceptor proteins are primarily based on the results of X-ray crystallography and spectroscopy studies. Of particular value are the observed band shifts in optical and vibrational spectra upon formation of the signaling (light-induced) state. However, the same set of experimental data has given rise to contradictory interpretations suggesting different structures of the dark and signaling states. To verify the specific mechanism of light-induced changes involving the rotation/tautomerization transformations with the conserved Gln residue near the flavin chromophore, we performed accurate quantum chemical calculations of the equilibrium structures, vibrational and absorption bands of the model systems mimicking the BLUF domain of flavoprotein AppA. Geometry optimization and calculations of vibrational frequencies were carried out with the QM(B3LYP/cc-pVDZ)/MM(AMBER) approach starting from the representative molecular dynamics (MD) snapshots. The MD simulations were initiated from the available crystal structures of the AppA protein. Calculations of the vertical excitation energies were performed with the scaled opposite spin configuration interaction with single substitutions SOS-CIS(D) method that enables efficient treatment of excited states in large molecular systems. The computed molecular structures as well as the spectral shifts (the red shift by 12÷16 nm in absorption and the downshift by 25 cm(-1) for the C4═O flavin vibrational mode) are in excellent agreement with the experimental results, lending a strong support to the mechanism proposed by Domratcheva et al. (Biophys. J. 2008, 94, 3872).

Light-Induced Structural Changes in a Photosynthetic Reaction Center Caught by Laue Diffraction

Photosynthetic reaction centers convert the energy content of light into a transmembrane potential difference and so provide the major pathway for energy input into the biosphere. We applied time-resolved Laue diffraction to study light-induced conformational changes in the photosynthetic reaction center complex of Blastochloris viridis. The side chain of TyrL162, which lies adjacent to the special pair of bacteriochlorophyll molecules that are photooxidized in the primary light conversion event of photosynthesis, was observed to move 1.3 angstroms closer to the special pair after photoactivation. Free energy calculations suggest that this movement results from the deprotonation of this conserved tyrosine residue and provides a mechanism for stabilizing the primary charge separation reactions of photosynthesis.

Blue Light Induces Global and Localized Conformational Changes in the Kinase Domain of Full-Length Phototropin

The blue-light photoreceptor phototropin plays a crucial role in optimizing photosynthesis in plants. In the two light-, oxygen-, or voltage-sensitive (LOV) domains of phototropin, the light stimulus is absorbed by the flavin chromophores. The signal is assumed to be transferred via dissociation and unfolding of a conserved J alpha helix element to the serine/threonine kinase domain. We investigated full-length phototropin from the green alga Chlamydomonas reinhardtii by Fourier transform infrared spectroscopy to shed light on the signal transfer within the protein and on the structural response of the kinase. Light-induced structural changes were assigned by comparing signals of the full-length protein with those of the truncated LOV1-LOV2-J alpha and LOV1-LOV2 and with those of deletion mutants. A loss of helicity originating from the J alpha linker helix was observed in LOV1-LOV2-J alpha in agreement with previous studies of LOV2-J alpha. Full-length phototropin showed reversible global conformational changes via several turn elements. These changes were suppressed in a deletion mutant lacking the J alpha linker and are attributed to the kinase domain. The loss of turn structure is interpreted as a light-induced opening of the kinase tertiary structure upon release of the LOV2 domain. Concomitant protonation changes of Asp or Glu residues in the kinase domain were not observed. A light-induced loss in helicity was observed only in the presence of a phototropin-characteristic 54-amino acid extension of the kinase activation loop, which is predicted to be located apart from the catalytic cleft. This response of the extension might play a significant role in the phototropin signaling process.

Light-Mediated Spatial Control via Photolabile Fluorescently Quenched Peptide Cassettes

Light-regulatable compounds are finding increasing utility as spatial and temporal probes of biological behavior. An independent measure of successful light-induced structural change is possible when alteration (e.g., activation, deactivation, etc.) of the bioprobe can be directly linked to a fluorescent readout. We have identified a series of photolabile fluorescently quenched cassettes that display extraordinarily large fluorescence enhancements upon photolysis. A pair of cassettes has been inserted into mitochondrial localization sequences to assess an organelle-targeted light-mediated release strategy for controlling biological activity. The peptide constructs are readily absorbed by mitochondria and subsequently can be cleaved in a light-dependent fashion as assessed by the predicted changes in absorbance and fluorescence.

Strong Hydrogen Bond between Glutamic Acid 46 and Chromophore Leads to the Intermediate Spectral Form and Excited State Proton Transfer in the Y42F Mutant of the Photoreceptor Photoactive Yellow Protein

In the Y42F mutant of photoactive yellow protein (PYP) the photoreceptor is in an equilibrium between two dark states, the yellow and intermediate spectral forms, absorbing at 457 and 390 nm, respectively. The nature of this equilibrium and the light-induced protonation and structural changes in the two spectral forms were characterized by transient absorption, fluorescence, FTIR, and pH indicator dye experiments. In the yellow form, the oxygen of the deprotonated p-hydroxycinnamoyl chromophore is linked by a strong low-barrier hydrogen bond to the protonated carboxyl group of Glu46 and by a weaker one to Thr50. Using FTIR, we find that the band due to the carbonyl of the protonated side chain of Glu46 is shifted from 1736 cm(-1) in wild type to 1724 cm(-1) in the yellow form of Y42F, implying a stronger hydrogen bond with the deprotonated chromophore in Y42F. The FTIR data suggest moreover that in the intermediate spectral form the chromophore is protonated and Glu46 deprotonated. Flash spectroscopy (50 ns-10 s) shows that the photocycles of the two forms are essentially the same except for a transition around 5 mus that has opposite signs in the two forms and is due to the chemical relaxation between the two dark states. The two cycles are coupled, likely by excited state proton transfer. The Y42F cycle differs from wild type by the occurrence of a new intermediate with protonated chromophore between the usual I(1) and I(2) intermediates which we call I(1)H (370 nm). Transient fluorescence measurements indicate that in I(1)H the chromophore retains the orientation it had in I(1). Transient proton uptake occurs with a time constant of 230 mus and a stoichiometry of 1. No proton uptake was associated however with the formation of the I(1)H intermediate and the relaxation of the yellow/intermediate equilibrium. These protonation changes of the chromophore thus occur intramolecularly. The chromophore-Glu46 hydrogen bond in Y42F is shorter than in wild type, since the adjacent chromophore-Y42 hydrogen bond is replaced by a longer one with Thr50. This facilitates proton transfer from Glu46 to the chromophore in the dark by lowering the barrier, leading to the protonation equilibrium and causing the rapid light-induced proton transfer which couples the cycles.

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

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 Pfr, 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 Pfr. In the biliverdin adduct, an additional component occurs that probably reflects a transition between two Meta-RC 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.

Different Role of the Jα Helix in the Light-Induced Activation of the LOV2 Domains in Various Phototropins

Phototropins (phot) are blue light receptors in plants which are involved in phototropism, stomatal opening, and chloroplast movements. Phototropin has two LOV domains (LOV1 and LOV2), and the LOV2 domain is responsible for activation of Ser/Thr kinase. There is an alpha-helix at the C-terminal side of the LOV2 domain, which is called the Jalpha helix. The functional importance of the Jalpha helix has been established for Arabidopsis phot1, where light-induced structural perturbation takes place in the Jalpha helix during the photocycle of LOV2 domains. However, the present FTIR study reports a different role of the Jalpha helix in light-induced signal transduction of LOV2 domains. Here we construct LOV2 domains with (LOV-Jalpha) and without (LOV-core) the Jalpha helix for Arabidopsis phot1 and phot2 and Adiantum neochrome 1 and compare their light-induced difference FTIR spectra. Light-induced protein structural changes differ significantly between LOV-Jalpha and LOV-core for Arabidopsis phot1 [Yamamoto, A., Iwata, T., Sato, Y., Matsuoka, D., Tokutomi, S., and Kandori, H. (2009) Biophys. J. 96, 2771-2778]. In contrast, the difference spectra are identical between LOV-Jalpha and LOV-core for Adiantum neochrome 1. In Arabidopsis phot2, the protein structural changes are intermediate between Arabidopsis phot1 and Adiantum neochrome 1. These results suggest that the conformational changes of the Jalpha helix and the interaction between the LOV-core and the Jalpha helix are different among phototropins. The role of the Jalpha helix for signal transduction in phototropins is discussed.