Unique Structural Relaxations and Molecular Conformations of

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Unique Structural Relaxations and Molecular Conformations of Porphyra-334 at the Excited State Makoto Hatakeyama, Kenichi Koizumi, Mauro Boero, Katsuyuki Nobusada, Hirokazu Hori, Taku Misonou, Takao Kobayashi, and Shinichiro Nakamura J. Phys. Chem. B, Just Accepted Manuscript • DOI: 10.1021/acs.jpcb.9b03744 • Publication Date (Web): 20 Aug 2019 Downloaded from pubs.acs.org on August 21, 2019

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The Journal of Physical Chemistry

Unique Structural Relaxations and Molecular Conformations of Porphyra-334 at the Excited State Makoto Hatakeyama,*ab Kenichi Koizumi,b Mauro Boero,e Katsuyuki Nobusada,cd Hirokazu Hori,f Taku Misonou,f Takao Kobayashi,g Shinichiro Nakamurabh a

Sanyo-Onoda City University, 1-1-1 Daigakudori, Sanyo-Onoda, Yamaguchi 756-0884, Japan

b

RIKEN Cluster for Science, Technology and Innovation Hub, 2-1 Hirosawa, Wako, Saitama 351-0198, Japan

Department of Theoretical and Computational Molecular Science, Institute for Molecular Science, Myodaiji, Okazaki, 444-8585, Japan c

d

Elements Strategy Initiative for Catalysts and Batteries (ESICB), Kyoto University, Katsura, Kyoto 615-8520, Japan

University of Strasbourg, CNRS, Institut de Physique et Chimie des Matériaux de Strasbourg UMR 7504, 23 rue du Loess, F-67034 Strasbourg, France e

f

Graduate School of University of Yamanashi 4-4-37, Takeda, Kofu, Yamanashi, 400-8510, Japan

Mitsubishi Chemical Corporation, MCC-Group Science and Technology Research Center Inc., 1000 Kamoshidacho, Aoba-ku, Yokohama 227-8502, Japan g

Computational Chemistry Applications Unit, Advanced Center for Computing and Communication, RIKEN, 2-1, Hirosawa, Wako, Saitama, 351-0198, Japan h

ABSTRACT:

Quantum chemistry based simulations were used to examine the excited state of

porphyra-334, one of the fundamental mycosporine-like amino acids present in a wide variety of aqueous organisms. Our calculations reveal three characteristic aspects of porphyra-334 related to either its ground or excited state. Specifically, (i) the ground state (S0) structure consists of a planar geometry in which three units can be identified, the central cyclohexene ring, the glycine- and the threonine-branch, reflecting the π conjugation of the system, (ii) the first singlet excited state (S1) shows a large oscillator strength and a typical ππ* excitation character and (iii) upon relaxation at S1 the originally ground-state planar structure undergoes a relaxation to a non-planar one, S1, especially at the carbon-nitrogen (CN) groups linking the cyclohexene ring to the glycine- or threonine-arm. The induced non-planarity can be ascribed to the fact that the carbon atoms of the CN groups prefer an sp3 hybridization in the S1 state. At the singlet state, these processes are unlikely to be trapped by singlet-triplet intersystem crossing especially when these occur in the hydrophilic zwitter-ion forms of porphyra-334. These results provide the missing information for the thorough interpretation of the stability of porphyra-334 upon UV irradiation.

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1. Introduction Porphyra-334 is an important member of the mycosporine-like amino acids (MMAs) family. It was originally identified in the red algae Porphyra tenera Kjellman1 and was subsequently found in other organisms.2,3 Porphyra-334, as well as other MMAs, is soluble in water, is characterized by a remarkable UV absorption, and exerts a photo-protective action exploited by a large variety of living organisms exposed to the sunlight. They exist within the marine macroalgae living in shallow water.4-10 Among MMAs, porphyra-334 is characterized by a large absorption band at 334 nm (ε = 4 × 104 M-1cm-1)1 and undergoes very little photodegradation (quantum yield ~ 2-4 × 104).11 This resumes to a remarkable photostability of porphyra-334 upon the solar irradiation. This photo-stability has been investigated experimentally to substance two major features: (i) a fast internal conversion from the excited state to the ground state11 and (ii) an efficient excess-energy dispersion upon a de-excitation to the surrounding environment as heat.12 The role of the internal conversion process has been probed by experiments on both porphyra-334 and other member of the MMA family.13,14 The excessenergy dispersion has been confirmed to be unaffected by the triplet states of porphyra334.11,12,15 In our former work, first principles molecular dynamics (FPMD) simulations on a hydrated porphyra-334 allowed us to disentangle the mechanism by which the excess energy is transferred from the photoexcited porphyra-334 to the surrounding water.16 Our results have shown that the efficient energy transfer is possible through the activation of the collective modes identified as hydrogen-bond (H-bond) stretching modes of the liquid water. Yet, the complete mechanism could not be fully captured by those simulations. What we

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missed was the relaxation process bringing porphyra-334 from its Franck-Condon excited state to its ground state. To fill this gap, here we report to quantum chemical simulations on the excited state of porphyra-334. To this aim, we focused on the zwitter-ionic and the less-polar forms of the neutral porphyra-334 (Figure 1) as the possible dominant forms over the charged forms (protonated/deprotonated, see supporting information) in an aqueous condition. In the present work, a thorough characterization of the spin-singlet excited state (S1) is done on the basis of a strict comparison of the calculated Franck-Condon excitation energy with the experimental absorption maximum. This analysis is complemented by the assessment of the optimized structures of the S1 state and the ground state S0. These analyses suggest the mechanism of a unique structural relaxation of porphyra-334 at the excited state. That is the large structural change of the whole molecular structure from the extensively π conjugated planar structure to the non-planar boat structure. Such a change originates from the cyclohexene part of porphyra-334 in which the sp3 and sp2 carbon atoms co-exist.17 Th S1 relaxation coordinate is associated with a possibility of fast internal conversion at porphyra-334. We also present how such a relaxation coordinate can be ascribed to the electronic states of porphyra-334.

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better accuracy of the calculation of the vertical excitation energies. Accounting for water solvation effects, the polarization continuum model (PCM) was adopted.21 Linear response formalism was used with PCM in investigating the excited states. Different possible initial structures were considered for the structural optimizations to include all the various orientations. Within these procedures, the terminal ligands can assume (-OH, -CH2OH, glycine-arm and threonine-arm of porphyra-334). These different initial structures were sampled utilizing molecular dynamics (MD) simulations within the density-functionaltight-binding (DFTB) approach on single porphyra-334 monomers.22 Following this procedure, ten independent configurations were sampled from the MD trajectories, which are sequential intervals of 10 ps simulations to pick up each geometry. For the structural optimizations of the excited states, we assumed the geometry of the FrankCondon state, namely the S0 optimized structure as the starting point. The steric configurations of the threonine-arm and the central cyclohexene ring were assumed to be in the L- and S-form, respectively (Figure 1) reflecting the experimental studies.23 The puckering of the central cyclohexene ring was taken into account in the preparation of the corresponding initial structures for MD simulations. 3. Results and Discussion 3.1. Analysis of Franck-Condon excitations and Comparison of absorption wavelength with experiment The first objective was to compute the Franck-Condon excited states corresponding to the system at the experimental absorption maximum (334 [nm] = 85.6 [kcal/mol]).1 For the optimized structure of the less-polar form A (Figure 1a) at the ground state (S0), the first spin-singlet excited state (S1) showed a large oscillator strength accompanied by the

Figure 1. Schematic representation of the four forms of porphyra-344 in the spin-singlet ground state (S0). The charge distributions of the zwitter-ion forms are based on the results of our calculation (see below). 2. Methods Simulations were done with the Gaussian 16 program package.18 Within the density functional theory (DFT) framework, the longrange corrected ωB97XD functional was used for almost all of the calculations.19 The longrange corrected functionals like ωB97XD are recommended in describing excited states having charge-transfer character. The parameter ω of ωB97XD functional was set to 0.2 [Bohr-1] in investigating the results shown in the text. Complementary calculations within the Symmetry Adapted ClusterConfiguration interaction (SAC-CI) method were done in some cases to refine the DFTbased results at a higher theoretical level.20 The localized 6-31G(d) basis sets was used to represent the electronic structure of all elements during the geometry optimization procedures. The basis set was then increased to the 6-311G(d,p) level aiming at achieving a 3



excitation energy of 104.1 kcal/mol at the TDDFT(ωB97XD) level from the sample showing the lowest S0 energy over other samples. This value, referred to the S1 excitation of the form A, reduces to 94.6 [kcal/mol] at the SAC-CI level, as reported in Table 1, and both are slightly larger than the experimental value (85.6 [kcal/mol]). An analogous result characterizes the less-polar form B (Table 1). At variance with these two cases, a better agreement with the experimental result was obtained for the S1 states of the zwitter-ion forms (C in Figure 1c and D in Figure 1d), and the agreement remarkably improves when the estimation is done at the SAC-CI level (86.3-87.2 [kcal/mol], see Table 1). Both the less-polar (A and B) and the zwitter-ion (C and D) forms show that the Franck-Condon S1 states have the large oscillator strength (i.e. non-dark state) and are mainly characterized by a ππ* excitation. For instance, in the case of the less-polar form A, the electron density difference (Δρ) between the S0 and S1 states (Δρ=ρ(S1)-ρ(S0), shown in Figure 2a) originates from the electronic excitation from the S0 ground state of the highest occupied π orbital on the CC double bond of the central cyclohexene ring (Figure 2b) to the lowest unoccupied π* orbital at the CNthr double bond of the threonine-arm in the S0 state (Figure 2c). These orbitals correspond to the ππ* electronic states. Thus, the S1 state of the lesspolar form A can be identified as the ππ* excited state. In the case of the zwitter-ion form C, the difference Δρ between the S0 and S1 states (Figure 2d) is mainly constituted by the lobes of the electronic distribution of the highest occupied π orbital located at the central cyclohexene ring (Figure 2e) and the lowest unoccupied π* orbital at the central ring, both in their S0 states (Figure 2f). The S1 state of the zwitter-ion form C can be identified as the ππ* excited state. The outcome of these Franck-Condon states

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analyses indicates that the experimental 85.6 [kcal/mol] excitation can be ascribed to the S1 excitation of the zwitter-ion forms. This ascription is consistent with another calculated result, which is the lower free energy of the zwitter-ion forms in the S0 state i.e. the state without the excitation than that of the less-polar forms in accordance with the water solvation effect with PCM. The free energies also indicate that the neutral forms are dominant than the charged forms (protonated/deprotonated, see SI) in an aqueous condition according to the equilibrium with water (e.g. equilibrium between “porphyra-334+H2O” and “[protonated porphyra-334]++OH-”). The free energies of these forms are summarized in SI. The S1 excitation energies reported in Table 1 refer to the neutral forms (less-polar and zwitter-ion) and have been obtained either at the TDDFT(ωB97XD) or at the SAC-CI theoretical level. The S1 excitation energies of the less-polar forms in the neutral charge state turned out to be nearly comparable with those of the deprotonated anionic forms having one C=N double bond. The S1 excitation energies of the neutral zwitterion forms are identical to those of the other deprotonated anion and protonated anion forms having no C=N double bond (see SI for details). Table 1. Excitation energies [kcal/mol] of the FrankCondon excited states for porpphyra-334. method

state

form A

form B

form C

form D

TDDFTa

S1

104.1

104.6

99.8

99.8

S2

121.2

117.2

125.4

123.2

S1

94.6

98.1

86.3

87.2

S2

114.4

111.7

136.8

140.8

SAC-CIa Exp.

85.6

Calculated values were obtained from the sample showing the lowest S0 state energy in each form. a


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the S0 state than that of the less-polar forms was also considered. Then, different possible orientations of the OCH3, CH2OH and COOH groups were taken into account by selecting ten configurations close to the S0 optimized structures. Assuming as starting configurations each one of the lowest energy structures in the S0 state, sketched in Figure 2, panels d-f, and Figure 3, panels a-c, one S1(ππ*) optimized structure could be obtained, and this is the one reported in Figure 3d-f. The S1(ππ*) optimized structure turns out to be characterized by three main structural features; (i) a stretching of the C-C bonds of the cyclohexene ring and of the C-N bonds of the threonine- and glycine-arms, (ii) a bending of the threonine- or glycine-arm from the central cyclohexene ring and (iii) the formation of a non-planar boat structure of the central cyclohexene ring (Figure 3e, f). In this respect, the S1(ππ*) optimized non-planar geometry is distinctly different from the S0 planar structure of Figure 3a-c. Such a nonplanar geometry implies that the π conjugation might be lost in the S1(ππ*) structure, resulting in a change of the ππ* character of the S1(ππ*) state. This issue will be detailed later in the ongoing discussion. In all the S1(ππ*) optimized structures, the bending of the threonine- or glycine-arm from the central ring turns out to be a constant characteristic independent from the initial arm orientation (Franck-Condon structure). Indeed, such a bending does not depend on whether or not the optimization reaches the S1 minimum before approaching the S1-S0 energy crossing point. Due to the bending of the boat form, the linking carbon atoms connecting the ring and the threonineor glycine arm, assumed a staggered conformation with the neighboring CH2 group (Figure 3e, f). From an energetical standpoint, the S1(ππ*) optimized structure (Figure 3d-f) shows an energy gap of 9.7

Figure 2. (a) Electron density difference (Δρ) between the S0 and S1 states of the less-polar form A, and (b) highest occupied π orbital of the S0 state for the less-polar form A. As in the other panels, the orbital population isosurface is shown at a value of 0.08 e/Å3. Panel (c) shows lowest unoccupied π* orbital of the S0 state for the less-polar form A, (d) the electron density difference Δρ between the S0 and S1 states of the zwitter-ion form C, (e) the highest π occupied orbital of the S0 state for the zwitter-ion form C, and, finally, (f) lowest unoccupied π* orbital of the S0 state for the zwitter-ion form C. The color code for the atoms is red for oxygen, blue for nitrogen, gray for carbon and cyan for hydrogen (in (b), (c) and (e), (f)), cyan for the decrease of electron density and orange for the increase of electron density (in (a) and (d)). 3.2. S1(ππ*) optimized structure To inspect the relaxation process of Porphyra-334 in the S1(ππ*) state, we performed structural optimizations in this excited state. To this aim, upon fixing the electronic excitation to S1, we considered the neutral zwitter-ion form C because of its better agreement with experiments in terms of Franck-Condon S1(ππ*) excitation energy. The lower energy of the zwitter-ion forms in 5



[kcal/mol] between the S1(ππ*) and S0 states at the TDDFT(ωB97XD) level.

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threonine-arm, one linking C atom between the ring and threonine arm, one C atom belonging to the CH2 group in the ring and the one C atom bonded to the OH and CH2OH groups (see ball-and-stick sketch of Figures 3a, b, d, e and the green-highlighted atom of Figure 3c, f). Thus, the threonine-arm bending can be measured by the Nthr-C-CH2C dihedral angle. Then, this configuration was analyzed via a partially constrained optimization at the S1(ππ*) state, being the constraint a set of different Nthr-C-CH2-C dihedral angles. The outcome of this analysis is summarized in Figure 4.

Figure 3. (a) S0 optimized structure of the neutral zwitter-ion form C, (b) side view of the S0 optimized structure, (c) schematic representation of the S0 optimized structure, (d) S1(ππ*) optimized structure and (e) side view of the S1(ππ*) optimized structure, and (f) schematic representation of S1(ππ*) optimized structure. The ball-and-stick atoms (a, b, d, e) and green-highlighted atoms (c, f) correspond to the heavy atoms of the Nthr-CCH2-C unit bended in the S1 optimized structure. 3.3. Relaxation energy along S1(ππ*) potential surface The energy levels of the S1(ππ*) and S0 states were monitored by looking at the threonineor glycine-arm bending. As mentioned above, the threonine- or glycine-arm bending is a constant feature in all the selected configurations used for the optimization of the S1(ππ*) state. We can then infer that these arm-bending motions represent a characteristic relaxation coordinate of the S1(ππ*) excited state. For instance, in the case of the conformation corresponding to the lowest energy in the S0 state, the threonine arm bends upon optimization of the S1(ππ*) state (Figure 3d-f). Such a bending is responsible for a change of the Nthr-C-CH2-C dihedral angle, defined by one N site of the

Figure 4. Total energies and orbital energies. The cyan point shows the S0 energy at the S0 optimized structure (S0/S0-opt), the pink one the S1(ππ*) energy at the S0 optimized structure (S1(ππ*)/S0-opt). Along the red dotted line, each circle shows the various S1(ππ*) energy at the (constrained) S1(ππ*) optimized structure (S1(ππ*)/S1-opt). Analogously, along the blue dotted line, the S0 energy at the (constrained) S1(ππ*) optimized structure (S0/S1-opt) is shown. For the orbital energies, the green color refers to the highest π orbital energy of the S0 state at the (constrained) S1(ππ*) optimized structure, and the orange one to lowest π* orbital energy of the S0 state at the (constrained) S1(ππ*) 6


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optimized structure). Total energies are relative to that of the S0/S0-opt point.

Such a trend is expected to be associated with porphyra-334 and its photostability. The S1-S0 energy gap reduction (Figure 4) was further examined in terms of the π and π* orbitals of the S0 state to get a deeper insight into the relationship among electronic state, molecular property and geometrical configuration. This analysis has shown that the highest π orbital of the S0 state (Figure 5) and the lowest π* orbital of the S0 state (Figure 6) are the major factor responsible for the S0 to S1 transition (Figure 2d-f). After the orbital analyses, we shall reconsider the ππ* character of the S1(ππ*) state based on the results at the non-planar S1(ππ*) optimized structure. 3.4. Change of highest π orbital The character of highest π orbital of the S0 state provides a rational basis to explain the S0 state energy increase. This orbital was inspected for both the S0 and S1(ππ*) optimized structures (Figure 5). In the first case, where the Nthr-C-CH2-C dihedral angle is about 210o, the highest π orbital of the S0 state is rather delocalized, extending from the glycine-arm to the threonine-arm through the central cyclohexene ring (Figure 5a, b). This wide distribution reflects the nature of the zwitter-ion form (see Figure 1); in fact, the π electron of the cyclohexene is partially shifted from the C-C bond of the cyclohexene ring to the CNthr or CNgly group through the intramolecular orbital interaction in the S0 state (Figure 1c, d). At the same time, at the S1(ππ*) optimized structure, corresponding to a Nthr-C-CH2-C dihedral angle of about 80o, the highest π orbital of the S0 state is rather localized on the cyclohexene ring (Figure 5c, d), as expected for a π electron localized on the C atoms of the carbon ring of the S1(ππ*) optimized structure. Such a localization reflects also the structural change from the planar S0 optimized structure (Figure 5a, b) to the non-planar S1(ππ*) optimized one (Figure

As sketched in Figure 4, the energy profiles of the S1(ππ*) excited state as a function of the Nthr-C-CH2-C dihedral angle show two trends. Namely, a sudden drop (pink asterisk in Figure 4) from the Franck-Condon (S1) excited state of the S0 optimized geometry to the dihedral S1(ππ*) optimized structure constrained (red line in the same figure), occurring at around 210o. The second trend, instead, is visible as a gradual decrease along the dihedral angle axis (red line in Figure 4) from ~210o to ~80o. The first sudden drop can be ascribed to the C-C and C-N bond stretching, reflecting the ππ* excitation character. The second trend, corresponding to a gradual decrease of the total energy of the S1(ππ*) state, is accompanied by an increase of the S0 one as a function of the Nthr-C-CH2-C dihedral change. The message Figure 4 conveys can then be summarized in a progressive decrease of the energy gap between the S1(ππ*) and S0 states as the S1(ππ*) relaxation proceeds along the dihedral axis. The decrease of the S1(ππ*) energy is stopped at a small Nthr-C-CH2-C dihedral angle (~ 70o), since the N atom of the bending threonine-arm approaches a H atom of the cyclohexene-ring (see Figure 3f) at such angle. The shapes of the S1(ππ*) and S0 energy curves (red and blue lines in Figure 4) are analogous to those of a case having “slopedtype” conical intersection (CI).24-26 Such a case shows the S1(ππ*) state minimum just before the region where the S1(ππ*) and S0 states are close to each other, being similar to the result of porphyra-334 (Figure 4). Thus, the corresponding energy curves can be confirmed to be analogous to each other, even though porphyra-334 is not yet investigated on its CI. The “sloped-type” CI does not branch the excited state relaxation pathway into multiple S0 state minima.25,26 7



5c, d), since this change hinders to various extents the intramolecular orbital interaction. The degree of localization of the π electron turns out to be consistent also with the energy increase of the highest π orbital and the energy increase of the S0 state due to the loss of the intramolecular orbital interaction.

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state and at the equilibrium dihedral angle Nthr-C-CH2-C ~ 210o, the lowest π* orbital is extended on both of two CN groups (Figure 6a, b). Conversely, at the S1(ππ*) optimized structure (Nthr-C-CH2-C ~ 80o), the lowest π* orbital is located exclusively on one CNthr group of the bended threonine-arm (Figure 6c, d) and consists of the 2p orbital of the C atom linking the cyclohexene ring to the threonine-arm. The major axis of this 2p orbital is tilted in such a way that it is not perpendicular to the neighboring C-C and CN bonds at the S1(ππ*) structure. By moving along the Nthr-C-CH2-C dihedral angle axis, the tilted 2p C orbital evolves gradually toward an sp3 hybridization, departing from the sp2 character of the linking C atom. The decrease in energy of the π* orbital along the Nthr-C-CH2-C dihedral occurs spontaneously because the sp3 hybridization prefers to assume a tilted alignment of the valence orbital lobes in the vicinity of the linking C atom to minimize both the Coulomb and Pauli repulsions. The reason why the π* orbital induces the sp3 hybridization will be clarified in the ongoing discussion.

Figure 5. (a) highest occupied π orbital of the S0 state in the S0 optimized structure, (b) side view of the highest occupied π orbital of the S0 state in the S0 optimized structure, (c) highest occupied π orbital of the S0 state in the S1(ππ*) optimized structure and (d) side view of the highest occupied π orbital. 3.5. Change of lowest π* orbital An analysis of the lowest π* orbital provides a key to understand the S1(ππ*) state energy decrease evidenced in Figure 4. For such a reason, this orbital was analyzed in both the S0 and the S1(ππ*) optimized structures (Figure 6). Focusing on the first one, in this S0 8 ACS Paragon Plus Environment

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the S1(ππ*) optimized structure, the π orbital is localized on the C-C bond of the cyclohexene ring, as shown in panels c and d of Figure 5, and this is different from the conjugating π orbital in the S0 optimized structure (Figure 5a, b). The different distributions of the π and π* orbitals, which are shown in the S1(ππ*) optimized structure as mentioned above, are also partially shown in the S0 optimized structure. This can be traced by looking at the electron density difference (Δρ) between the S0 and S1(ππ*) states (Figure 2d). Indeed, Δρ decreases in proximity of the central carbon of the cyclohexene ring (cyan-colored lobe in Figure 2d) and it increases at two linking C atoms connecting the ring to the arms (orange-colored lobes in Figure 2d). Such a distribution originates from an electron shift toward the protonated CNthr or CNgly groups in the S1(ππ*) state of the zwitter-ion form. Hence, the distribution of the π and π* orbitals can provide a hint to rationalize the structural change observed from the planar S0 optimized structure (Figure 3a-c) to the nonplanar S1(ππ*) optimized structure (Figure 3df). In fact, the S1(ππ*) excitation among these π and π* orbitals can weaken the π conjugation from the cyclohexene ring to the threonine-arm of porphyra-334. As a result, the S1(ππ*) excitation enables the cyclohexene ring to form the non-planar boat structure. The resulting geometric change to the boat structure induces the sp3 hybridization of the C atom linking the cyclohexene ring to the threonine-arm (Figure 6c, d). Moreover, the different distributions of the π and π* orbitals displace one electron from the C-C bond of the cyclohexene ring to the C-Nthr bond of the threonine-arm. This electron displacement, in turn, pushes the π electrons of the C-Nthr bond toward the N atom of this chemical bond. Then, this displaced π electron reverts to a component of the lone-pair electrons of the N atom. As a further check, we inspected

Figure 6. (a) lowest unoccupied π* orbital of the S0 state (= excited electron orbital of the S1(ππ*) state) in the S0 optimized structure, (b) side view of the lowest unoccupied π* orbital of the S0 state in the S0 optimized structure, (c) lowest unoccupied π* orbital of the S0 state in the S1(ππ*) optimized structure and (d) side view of the lowest unoccupied π* orbital. 3.6. Character of S1(ππ*) state and S1-S0 energy gap reduction The character of the S1(ππ*) state as ππ* holds also for the S1(ππ*) optimized structure (Nthr-C-CH2-C dihedral angle ~80o). * Nonetheless, the ππ character in the S1(ππ*) optimized structure is slightly different from that found in the S0 optimized structure (NthrC-CH2-C dihedral angle ~210o), as shown by the π and π* orbitals of Figures 5 and 6. This is not entirely unexpected, since the π and π* orbitals are localized far apart in the S1(ππ*) optimized structure, thus differing from those in the S0 optimized structure. Specifically, in 9



the atomic charges in the S1(ππ*) state, as reported in the SI, and their relative changes confirmed an increase in the electronic charge of the protonated CNthr group. A schematic representation of the type of chemical bonds and the electronic structure of porphyra-334 is shown in Figure 7 for the four optimized structures considered here ((a) So state at S0 optimized structure, (b) S1(ππ*) state at S0 optimized structure (FC state), (c) S0 state at S1(ππ*) optimized structure and (d) S1(ππ*) state at S1(ππ*) optimized structure). The non-bonding electron pairs of the N atoms are explicitly marked as a double dot in panels c and d. The S0 zwitter-ion form (Figure 7a) includes the deprotonated COO- group and the partially cationic C atoms connecting the cyclohexene ring to the glycine- and threonine-arms in a planar configuration. The S1(ππ*) zwitter-ion form (Figure 7b) carries a positive charge on the central C atom of the cyclohexene ring and this is the net effect of the π electron shift toward the linking carbon atoms. Along the S1(ππ*) relaxation pathway, the π electron shift weakens the π conjugation in the S0 state (Figure 7c) and is responsible for the boat structure of the cyclohexene ring in the S1(ππ*) state. Then resulting boat structure induces the sp3 hybridization of the C atom linking the cyclohexene ring to the threoninearm (Figure 7d).

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Figure 7. Schematic representation of the chemical bonding and electronic structure of porphyra-334. 3.7. Triplet states, hydration effects and photo-stability We discuss here on triplet states, hydration effects and the high photo-stability of porphyra-334. Experimentally, the hydrated porphyra-334 shows the photo-stability. Since the triplet states are more reactive than the singlet states, they can be origins of the decay. The hydration effects seem to be closely related to the suppression of possible intersystem crossing from the photo-excited spin-singlet ππ* state to the lower spin-triplet states. We must consider here two types of the triplet states: triplet ππ* and triplet nπ* states. First, we focus here on the crossing from the singlet ππ* state (S1 of porphyra-334) to the triplet nπ* state, since these states can have the large spin-orbit coupling as stated in ElSayed’s rules.27 The results of our calculations show that the zwitter-ion forms, labeled as C and D in Figure 1, are energetically more favorable than the less-polar forms (A and B in the same figure) in water, at least at the 10


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polarizable continuum model (PCM) level. Note that the zwitter-ion forms do not display the nitrogen lone-pair (n) orbitals on the CN groups, instead, they have N-H bonding orbitals. These zwitter-ion forms showed no triplet nπ* state in the energy region below 140 kcal/mol. Therefore, the zwitter-ion forms hardly have a chance to undergo the relaxation from the singlet ππ* state (S1) to the triplet nπ* state. By contrast, the less-polar forms (A and B of Figure 1), which are energetically more stable in dry conditions (vacuum) but less stable in hydrated condition, carry nitrogen lone-pair orbitals at the CNgly or CNthr group and possess a triplet nπ* state as the secondtriplet. In general, the less-polar form can have a risk to cross to its triplet state, in our case of porphyra-334 at the hydration, they are less stable than zwitter-ion forms. Second, we must consider the crossing from the photo-excited spin-singlet ππ* state (S1) to the triplet ππ* state. As shown in the energy profile (Figure S12 of SI), in our calculations, there is a triplet ππ* state (T1 of porphyra-334) below the singlet ππ* state (S1) regardless of the form (zwitter-ion or lesspolar), as is often the case in organic molecules having conjugated double bonds. Nonetheless, we can infer the possibility of less crossing to the triplet ππ* state. This is because, such a crossing should follow the small spin-orbit coupling as stated in ElSayed’s rule25 and it has to compete with the spin-allowed relaxation from the singlet ππ* state (S1) to the singlet ground state (S0). The spin-allowed relaxation will be faster than the crossing to the T1 state even in a situation where the S1 state becomes close to the T1 and S0 states energetically after the S1 relaxation (Figure S12). This is because, in such a situation, the S1 state will transit to the S0 state rapidly. However, a precise numerical evaluation will be a future subject.

4. Conclusions The present study provides a detailed insight into the intramolecular electronic state of porphyra-334 extending and complementing our former study, based on FPMD, of the energy release process occurring in a fully hydrated porphyra-334 molecule. We have shown that the structural relaxation at the S1(ππ*) excited state of porphyra-334 can significantly reduce the S1S0 energy gap by changing the planar S0 structure into the non-planar S1 configuration. The S1(ππ*) state conformation is the driving force responsible for the shift of the π electron of the cyclohexene ring toward the protonated CN group of the glycine- or threonine-arm in the Franck-Condon S1(ππ*) state. Such a π electron-shift triggers the sp3 hybridization at the carbon atom linking the cyclohexene ring to the glycine- or threonine-arm. Our quantum chemical simulations indicate that the zwitter-ion forms favor hydration without going through intersystem crossing in S1(ππ*) state, whereas the less-polar forms show a viable possibility to shift to a triplet state from the S1(ππ*) state. ASSOCIATED CONTENT Energies of the ground state and Franck-Condon excitation states at the neutral forms of porphyra-334. Energies of the ground state and Franck-Condon excitation states at the protonated-/deprotonated-forms. Mulliken’s atomic charges at the neutral forms. Energies of the spin-triplet states at the neutral forms. Coordinates of the neutral forms. This material is available free of charge via the Internet at http://pubs.acs.org.

AUTHOR INFORMATION Corresponding Author *Phone: +81-(0)836-39-6877. E-mail: [email protected]

Notes

Any additional relevant notes should be placed here.

ACKNOWLEDGMENT We dedicate this paper to the memory of our friend and colleague Katsuyuki Nobusada. We are deeply indebted to Prof. Shinji Saito at Institute for Molecular Science (Japan) for fruitful discussions and



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(14) Wooley, J. M.; Staniforth, M.; Horbury, M. D.; Richings, G. W.; Wills, M.; Stavros, V. G.; Unravelling the Photoprotection Properties of Mycosporine Amino Acid Motifs. J. Phys. Chem. Lett. 2018, 9, 3043-3048. (15) Inoue, Y.; Hori, H.; Sakurai, T.; Tokitomo, Y.; Saito, J.; Misonou, T. Measurement of Fluorescence Quantum Yield of Ultraviolet-Absorbing Substance Extracted from Red Alga: Porphyra yezoensis and its Photothermal Spectroscopy. Optical Review 2002, 9, 75-80. (16) Koizumi, K.; Hatakeyama, M.; Boero, M.; Nobusada, K.; Hori, H.; Misonou, T.; Nakamura, S. How Seaweeds Release the Excess Energy from Sunlight to Surrounding Sea Water. Phys. Chem. Chem. Phys. 2017, 19, 15745-15753. (17) Shishkina, S. V.; Shishkin, O. V.; Desenko, S. M.; Leszczynski, J.; Heterocyclic Analogues of Cyclohexene: Theoretical Studies of the Molecular Structures and RingInversion Processes. J. Phys. Chem. A 2007, 111, 2368-2375. (18) Frisch, M. J.; et al. Gaussian 16, Revision A. 03, Gaussian, Inc., Wallingford, CT, 2016. (19) Chai, J.-D.; Head-Gordon, M. Long-Range Corrected Hybrid Density Functionals with Damped Atom-Atom Dispersion Corrections. Phys. Chem. Chem. Phys. 2008, 10, 6615-6620. (20) Ehara, M.; Hasegawa, J.; Nakatsuji, H. Theory and Applications of Computational Chemistry: The First 40 Years, A Volume of Technical and Historical Perspectives, Elsevier, 2005, 1099-1141. (21) Tomashi, J.; Mennucci, B.; Cammi, R. QuJantum Mechanical Continuum Solvation Models. Chem Rev. 2005, 105, 2999-3093. (22) Frauenheim, T.; Seifert, G.; Elstner, M.; Hajnal, Z.; Jungnickel, G.; Porezag, D.; Suhai, S.; Scholz, R. A Self ‐ Consistent Charge Density ‐ Functional Based TightBinding Method for Predictive Materials Simulations in Physics, Chemistry and Biology. Physica Stat. Sol. B, 2000, 217, 41-62. (23) Klisch, M.; Richter, P.; Puchta, R.; Häder, D.-P.; Bauer, W. The Stereostructure of Porphyr-334: An Experimental and Calculational NMR Investigation. Evidence for an Efficient ‘Proton Sponge`. Helv. Chim. Acta, 2007, 90, 488-511. (24) Martinez, T.J. Seaming is Believing. Nature 2010, 467, 412413. (25) Boggio-Pasqua, M.; Robb, M. A.; Bearpark, M. J. Photostability via a Sloped Conical Intersection: A CASSCF and RASSCF Study of Pyracylene. J. Phys. Chem. A. 2005, 109, 8849-8856. (26) Hall, K. F.; Boggio-Pasqua, M.; Bearpark, M. J.; Robb, M. A. Photostability Via Sloped Conical Intersections: A Computational Study of the Excited States of the Naphthalene Radical Cation. J. Phys. Chem. A. 2006, 110, 13591-13599. (27) El Sayed, M. A. Spin-Orbit Coupling and the Radiationless Processes in Nitrogen Heterocyclics. J. Chem. Phys. 1963, 38, 2834-2838.

advices. This research was supported by JSPS KAKENHI (No. 25288012), Elements Strategy Initiative to Form Core Research Center (since 2012), and by MEXT, Japan. This research used computational resources of the FX100 system provided by Nagoya University through the HPCI System Research Project (project ID: hp190016). K. K. also thanks to the computer facilities of RCCS at Okazaki, Japan. Part of the calculations was performed on the HOKUSAI GreatWave system in RIKEN. M.B.~thanks Pôle HPC and Equipex Equip@Meso at the University of Strasbourg, and Grand Equipement National de Calcul Intensif (GENCI) under allocation DARI-A2 A0040906092.

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[TOC Graphic] We report a quantum chemical research on porphyra-334 how such a mycosporine-like amino acid exerts the great photo-protective action in various aqueous organisms.


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Unique Structural Relaxations and Molecular Conformations of

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