Research Article

Pathway for Mn-cluster oxidation by tyrosine-Z in the S2 state of photosystem II

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PNAS June 17, 2014 111 (24) 8723-8728; first published June 2, 2014; https://doi.org/10.1073/pnas.1401719111

  1. Edited by Pierre A. Joliot, Institut de Biologie Physico-Chimique, Paris, France, and approved April 24, 2014 (received for review January 28, 2014)

Significance

A key step in natural photosynthesis is the water-splitting reaction into molecular oxygen and hydrogen equivalents. Understanding the molecular mechanisms behind this photoreaction will unravel the secrets of solar energy conversion in biochemistry and may inspire the design of artificial biomimetic materials for green energy production. Photosynthetic water oxidation occurs in the Mn4Ca core of the photosystem II complex and proceeds through five subsequent steps S0 – S4 of the Kok cycle. Four electrons are sequentially removed from the Mn4Ca core by a nearby tyrosine, which is in turn oxidized by the photoactivated chlorophyll special pair. Using first principles multiscale atomistic simulations we clarify the thermodynamics and the kinetics for such electron abstraction in the S2 state.

Abstract

Water oxidation in photosynthetic organisms occurs through the five intermediate steps S0–S4 of the Kok cycle in the oxygen evolving complex of photosystem II (PSII). Along the catalytic cycle, four electrons are subsequently removed from the Mn4CaO5 core by the nearby tyrosine Tyr-Z, which is in turn oxidized by the chlorophyll special pair P680, the photo-induced primary donor in PSII. Recently, two Mn4CaO5 conformations, consistent with the S2 state (namely, S2A and S2B models) were suggested to exist, perhaps playing a different role within the S2-to-S3 transition. Here we report multiscale ab initio density functional theory plus U simulations revealing that upon such oxidation the relative thermodynamic stability of the two previously proposed geometries is reversed, the S2B state becoming the leading conformation. In this latter state a proton coupled electron transfer is spontaneously observed at ∼100 fs at room temperature dynamics. Upon oxidation, the Mn cluster, which is tightly electronically coupled along dynamics to the Tyr-Z tyrosyl group, releases a proton from the nearby W1 water molecule to the close Asp-61 on the femtosecond timescale, thus undergoing a conformational transition increasing the available space for the subsequent coordination of an additional water molecule. The results can help to rationalize previous spectroscopic experiments and confirm, for the first time to our knowledge, that the water-splitting reaction has to proceed through the S2B conformation, providing the basis for a structural model of the S3 state.

For 2.5 Gy photosynthetic organisms have used the photosystem II complex (PSII) to capture light energy from the sun and convert it into chemical energy stored within energy-rich carbohydrates (1). The water oxidation reaction, occurring in the reaction center of PSII, represents the central step of the natural photosynthetic process, leading to the formation of molecular oxygen and hydrogen equivalents. A deep understanding of the photosynthetic water-splitting mechanism may serve as a valuable source of inspiration for the development of artificial devices able to store solar energy in environmentally friendly fuels, such as molecular hydrogen (2⇓⇓⇓–6). The active site of the PSII enzyme, where the water-splitting reaction takes place, consists of a core of four Mn ions and one Ca ion connected together through μ-oxo bridges in a cubane-like aggregate (7). Water oxidation proceeds through five sequential S0–S4 steps known as the Kok cycle (8). At each step of the catalytic cycle the absorption of photons turns out the oxidation of the tyrosyl group of a nearby tyrosine (i.e., Tyr161, also known as Tyr-Z in the D1 subunit of PSII), which acts as an intermediate in the electron transfer between the Mn4CaO5 cluster and the primary donor P680 (9, 10).

The molecular structure for the different states of the Kok cycle was largely investigated in the past decades by extended X-ray absorption fine structure experiments (11⇓⇓–14) and X-ray crystallography (15⇓–17), thus revealing atomic details at an increasing level of accuracy (7). In parallel the electronic and magnetic properties, characterizing steps of the catalytic cycle, were investigated by EPR experiments (18⇓⇓⇓⇓⇓–24), with particular attention to the well-characterized S2 state (25). The combination of this large number of experimental results within a computational modeling framework gave the opportunity to understand some of the atomic details underlying the water-splitting reaction in the oxygen-evolving complex (OEC) (26⇓⇓⇓⇓⇓⇓⇓⇓–35). Recently, Pantazis et al. (36) have suggested the presence of two possible interconvertible structures representative of the S2 state. The two models, namely, the S2A state, characterized by a S = 1/2 spin ground state, and the S2B state, characterized by a S = 5/2 spin ground state (Fig. 1), can explain the presence of the two distinct EPR signals revealed at cryogenic temperatures (i.e., a multiline signal indicative of a ground state characterized by a spin S = 1/2 and a second signal at g = 4.1 consistent with a spin S = 5/2). In a recent work (37) we characterized by extensive quantum mechanics/molecular mechanics (QM/MM) ab initio simulations the free-energy profiles for the interconversion between the two above-mentioned conformations, suggesting that the transition from the S2 to S3 state should proceed passing first by the S2A and subsequently through the S2B state. Still, clear evidence confirming such a hypothesis is missing.

Fig. 1.
Fig. 1.

Ab initio QM/MM model of photosystem II. (Right) The QM region, consisting of 224 atoms, is shown in balls and sticks representation. (Upper Left) A selection of the the most important residues and distances involved in the oxidation of the Mn4CaO5 cluster by the radical Tyr-Z are sketched. (Lower Left) Representation of the two investigated conformations S2A and S2B.

Here, using the same approach previously adopted, we characterized different (spin) energy surfaces along the interconversion path between the S2A and S2B states after the removal of one electron from the QM region (Fig. 1). QM/MM molecular dynamic simulations were additionally carried out for the two models in their respective spin ground state: the low spin (LS) state for the S2A model and the high spin (HS) state for the S2B model. The present results show for the first time, to our knowledge, the occurrence of a proton coupled electron transfer (PCET) in the S2B state resulting in the oxidation of the Mn4 ion by the tyrosyl group of Tyr-Z.

Results and Discussion

In a recent work (37) we carried out thermodynamic integration calculations to determine the relative stability of the two states S2A and S2B and to estimate the free energy barrier for their interconversion. Here, to investigate whether the removal of one electron influences the thermodynamic and the kinetic of such conformational change, we calculated by thermodynamic integration the free energy profiles for the respective oxidized states along the reaction coordinate ξ (defined as the difference between Mn4-O5 and Mn1-O5 distances, see also Method section and Supplementary Information). In the following, consistently with the notation adopted by Pantazis et al. (36), we will refer to the two conformers representative of the oxidized S2 state as [S2A]+ and [S2B]+ and to the two reduced ones as [S2A]0 and [S2B]0. If not specified S2A and S2B will simply refer to the respective geometrical configurations. It should be pointed out that the notation [S2]+ may refer to different oxidation patterns, such for instance [Mn4(IV)/Tyr-Z] or [Mn(III)Mn3(IV)/Tyr-Z•].

Free-Energy Landscape of the [S2]+ State.

The free-energy profiles calculated for the oxidized system [S2]+ in both LS (Sz = 0, red solid line) and HS (Sz = 3, green solid line) states are reported in Fig. 2, Bottom. The LS profile does not significantly differ with respect to that obtained for the reduced state (dashed red line) (37). In both LS cases the transition from the S2A state to the S2B state is slightly endergonic with similar energy barriers (∼9.9 kcal/mol in the oxidized system and ∼10.6 kcal/mol in the reduced system).

Fig. 2.
Fig. 2.

Electronic structure and energetics along the interconversion between the two S2 structural models upon removal of one electron from the system ([S2]+ oxidized state). The two upper panels follow changes in the spin population (absolute value) of the Mn ions (left scale) and of the Tyr-Z (right scale) as a function of the reaction coordinate ξ for the two models. In the bottom panel, the calculated free energy profiles (solid lines) show that the HS ground state in [S2B]+ is thermodynamically favored by 2.6 kcal/mol with respect to the LS [S2A]+ state. The relative stability in the oxidized system is therefore inverted compared with the reduced [S2]0 state (dashed lines; data from ref. 37).

The removal of one electron in the LS configuration is therefore not affecting the relative stability of the two states, confirming the larger stability of the S2A state with respect to the S2B state, independently on the overall oxidation state of the system. Conversely, the HS surface for the oxidized state (solid green line) shows a qualitatively different profile, indicating a significant exergonicity of the transition from S2A+ to S2B+. Here, the [S2B]+ state shows a larger stability with respect to the [S2A]+ state (ΔG≃−2.6 kcal/mol). Moreover, the transition state seems to correspond to a shorter value of the reaction coordinate ξ with respect to the LS case. The free energy profiles indicate that the transition between [S2A]+ and [S2B]+ occurs on the LS surface, as in the case of the [S2A]0-to-[S2B]0 transition (37). Thereafter, the conversion from LS to HS state, occurring in the [S2B]+ state, is characterized by a decrease of the free energy, inverting the thermodynamic stability of the two conformers.

To understand which are the changes in the electronic structure occurring along the transition, we reported in Fig. 2, Top and Middle the spin populations of the four Mn ions and the tyrosyl group of Tyr-Z as a function of the reaction coordinate ξ in the oxidized system for both the LS and HS configurations. In the LS state the spin population inversion of the two Mn1 and Mn4 ions occurs, consistent with a transition from the Mn1(III)Mn4(IV) to the Mn1(IV)Mn4(III) state. Conversely, Tyr-Z keeps a radical nature for all of the simulated values of the reaction coordinate. It has to be pointed out that the Mulliken spin populations, although very useful to follow the evolution of the electronic structure along dynamics, should not be taken strictly quantitatively. The same behavior for the Mn1 and Mn4 ions was already reported for the LS state in the reduced system (37). Similarly to the LS case, in the HS state the spin populations of Mn1 and Mn4 undergo a concerted inversion in the proximity of the transition state (ξ ≃ −0.1 Å), with the residue Tyr-Z still characterized by a radical state. Notably, for values of the reaction coordinate close to the typical value characterizing the S2B state, the spin population of Mn4 exhibits a clear transition concerted with a spin state transition of the Tyr-Z. In particular our results show that for geometries close to the S2B state, Tyr-Z loses its radical nature with a simultaneous transition of the spin population of Mn4 consistent with a formal oxidation number of IV. When the system is in the HS state, an electron transfer between the Mn4 and Tyr-Z in a S2B-like configuration is therefore occurring during our constrained QM/MM simulations.

PCET.

To better investigate the observed electron transfer and to characterize the electronic states in [S2A]+ and [S2B]+ we carried out QM/MM calculations with no constraints applied on the reaction coordinates by removing one electron from the system starting from the [S2A]0 and [S2B]0 states. The two QM/MM simulations for the two oxidized states [S2A]+ and [S2B]+ were carried out in their respective spin ground state (i.e., Sz = 0 for [S2A]+ and Sz = 3 for [S2B]+). The Mulliken spin populations of the four Mn ions and the Tyr-Z were monitored over the simulation time. In the [S2A]+ state (Fig. 3, Top Left) the starting spin populations of the four Mn ions remain constant during the simulation time, thus keeping their oxidation states consistent with the typical S2 state of the Kok cycle [Mn(III)Mn3(IV)]. The removal of the electron from the system mainly affects the electronic structure of Tyr-Z, showing a clear radical nature along most of the simulation time. The (time-dependent) protonation states of the two titratable residues Tyr-Z and His190 were also monitored (Fig. 3, Middle Left), reporting both the distances between the tyrosyl oxygen of Tyr-Z and the respective titratable hydrogen atom and between the imidazole nitrogen of His190 and the above-mentioned hydrogen atom (see also Fig. 1, where the two above-mentioned distances are defined as d1 and d2, respectively). After the first 0.5 ps, characterized by mutual proton exchanges between the two titratable residues, the proton was found to bind the His190, thus leaving Tyr-Z in its deprotonated (radical) state. It has to be pointed out that the protonation state of Tyr-Z and the spin density of the tyrosyl group show a strong correlation, being the radical state stabilized by the removal of the proton. Distances between a hydrogen atom belonging to the water molecule W1 and both the water oxygen of W1 and the carboxylic oxygen of Asp61 (see Fig. 1, where the two above mentioned distances are defined as d3 and d4, respectively) were calculated as a function of simulation time for the state [S2A]+ and shown in Fig. 3, Bottom Left. The reported results show the absence of proton exchanges between the titratable residue Asp61 and the water molecule W1 despite the fact that a transient hydrogen bond between the (donor) W1 molecule and the (acceptor) Asp61 is present during the simulation, in particular after the first 0.5 ps. Similar to the case of the [S2A]+ state, the removal of the electron from the system in the S2B configuration leads to a radicalization of the tyrosyl group of Tyr-Z in the first steps (0–0.2 ps) of the unconstrained QM/MM simulation (Fig. 3, Top Right). In the same time range the four Mn ions of the OEC cluster keep their starting spin populations consistent with their expected oxidation state in the S2 state of the Kok cycle. Nevertheless, after 0.2 ps of simulation, the spin populations of Tyr-Z and Mn4 undergo a drastic transition, leading, after 0.6 ps of simulation, to a stable singlet state for the Tyr-Z and to a simultaneous oxidation of the Mn4 ion, from Mn(III) to Mn(IV). The concerted spin transition between the tyrosyl group and the Mn4 ion gives strong evidence for the presence of an electron transfer from the Mn4 ion to the tyrosyl radical of the Tyr-Z residue. The electron transfer event converts the overall electronic oxidation state of the Mn4CaO5 cluster to the electronic configuration suggested for the S3 state of the Kok cycle [i.e., Mn1(IV)Mn2(IV)Mn3(IV)Mn4(IV)] (38). Moreover, this transition is solely observed in the [S2B]+ state, whereas in the [S2A]+ state the Mn ion with the lower oxidation state [Mn1(III)] does not show any spin transition during the simulation time.

Fig. 3.
Fig. 3.

QM/MM molecular dynamics in the LS [S2A]+ (Left) and HS [S2B]+ (Right) state. [S2A]+ state: In the top panel the spin populations of the four Mn ions (left scale) and the Tyr-Z (violet line, right scale) show that in the simulated time the removed electron remains localized on the Tyr-Z radical. Selected distances (as sketched in Fig. 1) are reported in the middle and bottom panel showing that along dynamics His190 remains protonated, whereas Asp61 is negatively charged. [S2B]+ state: In the top panel the changes in spin population of the four Mn ions (right scale) and the Tyr-Z (violet line, left scale) are reported as a function of the simulated time. A spontaneous Mn cluster oxidation by the radical Tyr-Z in the [S2B]+ state occurs through a proton coupled electron transfer mechanism on sub-picosecond timescale (the electron has been transferred from Mn4 to Tyr-Z). Middle and bottom panels report the analysis of selected distances showing the occurrence of a proton transfer from the water molecule W1 to Asp61 and the prevalent neutrality of His190 after the Tyr-Z electron transfer.

Intriguingly, the electron transfer was found to occur in parallel with the proton exchange between Asp61 and the water molecule W1 (Fig. 3, Bottom Right), thus giving rise to the concurrent protonation of Asp61 and the formation of a hydroxide ion in place of the W1 water molecule as ligand of Mn4. Furthermore, the rearrangement of the OEC electronic structure was shown to stabilize the protonation state of both Tyr-Z and His190 residues (Fig. 3, Middle Right). Indeed, after 0.6 ps of simulation, the proton, initially hopping between these two amino acids, stably binds the tyrosyl oxygen, in contrast with the case of the [S2A]+ state. The apparent mutual influence between the protonation states of Asp61 and the residues pair Tyr-Z/His190 may have originated from the hydrogen-bonding network connecting Asp61 and His190 (39). Four snapshots representative of the events occurring during the unconstrained QM/MM dynamic were extracted from [S2B]+ state trajectory and shown in Fig. 4 with the respective spin density iso-surfaces.

Fig. 4.
Fig. 4.

Spin densities of selected snapshots along the [S2B]+ dynamics. Representative snapshots were chosen to describe the PCET steps. Blue and red isosurfaces refer to the down and up spin densities, respectively. To improve the visual representation of the spin densities a different density cutoff has been used for the Tyr-Z and the Mn cluster.

To the best of our knowledge the presented results give for the first time evidence that the transition through the S3 state proceeds from the S2B conformation, as suggested in recent studies (37, 40). Moreover the calculated free energy profiles indicate the [S2A]+-to-[S2B]+ transition half-time is in the microsecond timescale (on the basis of the Eyring–Polanyi equation). In this respect, the rate-limiting step observed for the S2→S3 transition (41) could be ascribed to the conversion from the S2A to the S2B structure.

Conformational Changes Along the S2-to-S3 Transition.

The electronic and protonation changes occurring into the Mn4CaO5 cluster in the [S2B]+ state have important consequences on the geometrical properties of the cluster itself. In particular, we have monitored the Mn1–Mn4 distance, because it was already suggested that an increase of such distance could facilitate the coordination of an additional water molecule to the Mn4 ion (37). The Mn1–Mn4 distance distributions for both the [S2A]+ and [S2B]+ QM/MM simulations were compared with the respective reduced states (Fig. S1). The oxidation of the system clearly leads to an increase of the Mn1–Mn4 distance for both the investigated conformations. Nevertheless, the effect is much more prominent in the S2B state with respect to the S2A state. In particular, for the oxidized state [S2B]+ conformations characterized by a distance larger than 5.4 Å were found to be notably populated. The analysis of the conformations sampled during the dynamics also reveals that the proton transfer is followed by a rotation of 120° of the Asp61 side chain dihedral angle (Fig. 3, Bottom Right), thus leaving the W1 water molecule in a stable deprotonated state (i.e., hydroxide ion) strongly coordinated to Mn4.

The proton shuffling between the water molecule W1 and the Asp61 could have important implications on the water-splitting mechanism. In particular, two possible scenarios can be supposed. In the first this proton transfer corresponds to the initial step of a cascade mechanism, finally pumping the proton across the thylakoid membrane into the lumen. In this view, as previously suggested (42⇓–44), Asp61 can be regarded as a gate for the proton channel.

In the second scenario the proton originally bound to water molecule W1 never leaves the vicinity of the Mn4CaO5 cluster, just shuffling between Asp61 and water W1 depending on the oxidation state of the system. In this respect, we can suggest that the proton transfer occurring from the water to the aspartic acid, leaving a net negative charge (i.e., the hydroxyl group) directly coordinated to the Mn4 ion, could trigger the oxidation of the Mn4 by the radical Tyr-Z. A similar mechanism was already suggested in respect to the Tyr-Z/HisZ moiety with the proton moving back and forth between Tyr-Z and His190 (10), thus catalyzing the deprotonation reaction on the Mn4CaO5 cluster. Similarly, the proton shuffling between W1 and Asp61 could catalyze the oxidation of the Mn cluster.

Conclusions

The mechanism of the Mn cluster oxidation by Tyr-Z in the S2 state of PSII was investigated by exploring the free-energy landscape of the oxidized S2 state (i.e., [S2]+) around the two conformers S2A (having an LS ground state) and S2B (having an HS ground state). The presented results can provide a detailed picture of the intermediate states along the S2-to-S3 transition, as summarized in Fig. 5. Before the oxidation of the Tyr-Z, in the S2 state, the [S2A]0 conformer is the most stable from the thermodynamic point of view (37), and the two conformers are separated by a free-energy barrier of about ∼10.6 kcal/mol, corresponding to a half-time τ1/2(A→B) in the microsecond timescale at room temperature. Upon oxidation, the thermodynamic stability is inverted, the [S2B]+ state becoming significantly more stable, as schematized in the second equilibrium of Fig. 5. In the [S2A]+ state, the electron hole is localized on Tyr-Z, which has a radical character and donates its titratable proton to the nearby His190. This radical/ionic pair remained stable along the 6 ps of the simulated dynamics.

Fig. 5.
Fig. 5.

Sketch of the proposed intermediates in the transition between S2 to S3 states of the Kok cycle.

In the [S2B]+ state the Tyr-Z radical is not stable anymore, and an electron transfer takes place on a sub-picosecond timescale, leading from a Mn(III)Mn3(IV) to a Mn4(IV) metal cluster. Simultaneously, a proton transfer is occurring at the Mn cluster site, namely from W1 to Asp61, which becomes neutral. This scheme implies that in the S2B conformation, characterized by an HS ground state (SI Materials and Methods and Table S1), an electron transfer occurs spontaneously from the Mn4(III) ion to the radical Tyr-Z, whereas in the S2A conformation (characterized by an LS ground state) the radical Tyr-Z seems to be stable in the simulation time. Interestingly, low-temperature EPR experiments have reported the so-called split signal (45⇓–47) during the S2-to-S3 transition, which is likely generated by the coupling between the Tyr-Z radical (S = 1/2) and an LS (S = 1/2) located on the Mn cluster (48, 49). This signal can be therefore compatible with the [S2A]+ state here described, in which the radical tyrosyl group is coupled with the Mn cluster in an LS state. However, to the best of our knowledge, no EPR signals have been reported so far that might correspond to a tyrosyl radical coupled with an HS state. From our calculations this state would correspond to the [S2B]+ configuration with the radical Tyr-Z and the Mn cluster in the Mn(III)Mn3(IV) oxidation state. However, this state is shown to be highly unstable, spontaneously leading on a sub-picosecond timescale to the reduction of the Tyr-Z by the Mn cluster reaching the Mn4(IV) oxidation state (Fig. 3, Top Right) and therefore not detectable even at the low temperatures usually achieved by the EPR experiments.

The change of relative stability upon oxidation also provides us with strong evidence that the S2A state precedes the S2B state during the S2-to-S3 transition, as already suggested in previous papers (40, 50) and supported by recent calculations (37). The picture emerging from our data also suggests that the thermally activated passage from [S2A]+ to [S2B]+ conformer may represent the rate-determining step for the S2-to-S3 transition, albeit the subsequent water binding may also contribute to such a step. The estimated free-energy barrier for the S2-to-S3 transition, although it cannot be taken with high precision owing to the intrinsic limitations of the density functional theory plus U (DFT+U) approach and to the statistical error owing to finite time of simulations, is indeed compatible with the fact that the observed time for the electron transfer from the Mn cluster to the radical Tyr-Z occurs in the microsecond range (51) upon light excitation. The electron rearrangement significantly affects the metal–metal distances of the Mn cluster. In particular, the resulting Mn4(IV) geometry has an elongated Mn1–Mn4 distance, conferring to the cluster a more open structure, which might lead to an increase in the molecular volume. Interestingly, a significant increase in the molecular volume has been also observed by Dau and coworkers in photo-acoustic experiments between S2 and S3 states (52, 53) and attributed to a proton release from the cluster. In this regard it should be noted that the PCET mechanism involving the simultaneous proton shuffling between Tyr-Z and His190 and the electron transfer from the Mn4 to the radical Tyr-Z cannot be associated with the proton release observed by Dau and coworkers (52). Moreover, because in our model system the proton suggested by Dau and coworkers to be released in 30 μs might already be absent, the subsequent half-time of 180 μs observed for the Tyr-Z rereduction (54) could be ascribed (at least partially) to the interconversion between the S2A and the S2B state.

The increase of solvent exposure shown by the [S2B]+ model might promote the binding of one additional water molecule and the formation of the hexa-coordinated Mn4, as already suggested by Cox and Messinger (40). The binding of an external water molecule is consistent with mass spectrometric results, although these data exclude the possibility that this water is the substrate (both substrate water molecules are already bound in the S2 state) (40). Based on our calculations the water binding is likely to occur between the [S2B]+ and S3 states, as sketched in Fig. 5. Such hexa-coordinated S2B geometry has been already observed by us in a previous simulation (37), and a similar binding mechanism might be expected in [S2B]+. These results also indicate that the S2B model should be taken as a reference point for modeling the water oxidation reaction, at variance with what has been considered so far in electronic structure calculations (55). In summary, the present work provides structural insights on the conformational changes occurring in the Kok cycle between the S2 and S3 states, thus helping to rationalize previously suggested water-splitting mechanisms (56, 57). The data clearly show that the S2A structure has to precede the S2B state along the cycle, providing a structural basis of the S3 state.

Materials and Methods

Model Definition.

Starting from a classical simulation of a PSII dimeric complex embedded in a membrane bilayer, we built our QM/MM model by extracting a subset of about 40,000 atoms including all of the amino acids of the D1, D2, and CP43 polypeptide chains, the neighboring cofactors, and the water molecules present in the structure (details reported in ref. 58). The considered quantum region is composed by the metal core, its ligands (Asp170, Glu189, His332, Glu333, Asp342, Ala344, and CP43-Glu354), their closest residues (Asp61, Tyr161, His190, His337, Ser169, and CP43-Arg357), the 10 closest water molecules plus the four water/hydroxide molecules coordinated to the Mn4CaO5 cluster, and the chloride anion next to Glu333.

Computational Procedure.

All QM/MM calculations were carried out using the CP2K package (59, 60). The MM system is treated at a classical level by the general AMBER force field (GAFF) (61) for the cofactors and AMBER99SB force field (62) for the protein residues, as detailed in our previous studies (37, 58). To treat the electronic structure of the quantum region we used the spin-unrestricted Kohn–Sham DFT+U scheme (63⇓–65) using the functional Perdew–Burke–Ernzerhof (66) with U=1.16 eV for the Hubbard correction (58). For all atoms we used Goedecker–Teter–Hutter pseudopotentials (67, 68). We adopted the Gaussian/plane-wave scheme (60) as implemented in the CP2K package, using the DZVP-MOLOPT-SR-GTH Gaussian basis set optimized for molecular systems (69) and a 380 Rydberg cutoff for the plane-wave basis set. QM/MM Born–Oppenheimer dynamics were performed with a time step of 0.5 fs in NVT ensemble with T = 298 K. Free-energy calculations were carried out on different spin surfaces by thermodynamic integration (70) using the difference of distances ξ=d(Mn4,O5)−d(Mn1,O5) as reaction coordinate for the transition between the S2A and S2B structures of the Mn4CaO5 cluster. The force sampling was performed on 31 points per pathway with ξ=−1.5,−1.4,…,+1.5 Å, averaging for each point the Lagrange multipliers on 1.0 ps of QM/MM dynamics, after an equilibration of ∼1.0 ps. The energy gap in [S2A]+ structure between the LS and HS states reported in Fig. 2 was estimated by QM/MM geometry optimization. The QM/MM molecular dynamics of the [S2A]+ and [S2B]+ states were carried out for a total computed time of 6.0 ps each.

Acknowledgments

We thank Holger Dau, Michael Haumann, and Ivelina Zaharieva for useful discussions. Funding was provided by the European Research Council Project 240624 within the VII Framework Program of the European Union. Computational resources were supplied by CINECA, PRACE infrastructure, and the Caliban-HPC center at the University of L’Aquila.

Footnotes

  • ↵1D.N. and D.B. contributed equally to this work.

  • ↵2To whom correspondence should be addressed. E-mail: l.guidoni{at}gmail.com.

  • Author contributions: D.N., D.B., and L.G. designed research; D.N. and D.B. performed research; D.N., D.B., and L.G. analyzed data; and D.N., D.B., and L.G. wrote the paper.

  • The authors declare no conflict of interest.

  • This article is a PNAS Direct Submission.

  • This article contains supporting information online at www.pnas.org/lookup/suppl/doi:10.1073/pnas.1401719111/-/DCSupplemental.

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Pathway for Mn-cluster oxidation by Tyr-Z in PSII
Daniele Narzi, Daniele Bovi, Leonardo Guidoni
Proceedings of the National Academy of Sciences Jun 2014, 111 (24) 8723-8728; DOI: 10.1073/pnas.1401719111
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