Cationic State Distributions over Chlorophyll Pairs in Photosystem I and II

Photosystem I (PSI) and II (PSII) possess chlorophyll pairs P~A~/P~B~ and P~D1~/P~D2~, respectively. These chlorophylls are the primary electron donors in the light-induced electron transfer. After the electron transfer, the radical cation remains on these chlorophyll pairs, forming [P~A~/P~B~]^·+^ and [P~D1~/P~D2~]^·+^. The positive charge distributions over the two chlorophylls were reported to be 10/90-50/50 for P~A~^·+^/P~B~^·+^ [1,2] and 70/30-80/20 for P~D1~^·+^/P~D2~^·+^ [3,4]. To clarify the origin of the distributions, we calculated ratios of P~A~^·+^/P~B~^·+^ and P~D1~^·+^/P~D2~^·+^ with a quantum mechanical/molecular mechanical (QM/MM) approach and the redox potentials (_E_~m~) of monomeric chlorophylls P~A~, P~B~, P~D1~, and P~D2~ with an electrostatic continuum-model approach, using the crystal structures of PSI [5] and PSII [6]. 
1) Our QM/MM calculation reproduced the experimentally measured ratios of P~A~^·+^/P~B~^·+^ [1,2] and P~D1~^·+^/P~D2~^·+^ [3,4]. The calculated ratios were strongly correlated with the calculated _E_~m~ values. 2) We analyzed residues on puseudo-symmetrical subunit pairs PsaA/PasB and D1/D2 that shifted _E_~m~ of P~A~, P~B~, P~D1~, and P~D2~ and identified the residue pairs responsible for the P~A~^·+^/P~B~^·+^ and the P~D1~^·+^/P~D2~^·+^ ratios. In PSII, the difference in the electrostatic protein environments between D1 and D2 was significant in determining the P~D1~^·+^/P~D2~^·+^ ratio, whereas geometric differences between P~A~ and P~B~ (P~A~ as the C13^2^ epimer of chlorophyll a and the H-bond pattern) played a role in determining the P~A~^·+^/P~B~^·+^ ratio in PSI.

References:
[1] A. N. Webber, W. Lupitz, _Biochim. Biophys. Acta. 1507_ (2001) 61.
[2] M. Pantelidou, P. R. Chitnis et al., _Biochemistry 43_ (2004) 8380.
[3] I. H. Davis, P. Heathcote et al., _Biochim. Biophys. Acta. 1143_ (1993) 183.
[4] T. Okubo, T. Tomo et al., _Biochemistry 46_ (2007) 4390.
[5] P. Jordan, P. Fromme et al., _Nature 411_ (2001) 909.
[6] Y. Umena, K. Kawakami et al., _Nature 473_ (2011) 55. 
&#xa

Tyrosine Deprotonation and Associated Hydrogen Bond Rearrangements in a Photosynthetic Reaction Center

Photosynthetic reaction centers from Blastochloris viridis possess Tyr-L162 located mid-way between the special pair chlorophyll (P) and the heme (heme3). While mutation of the tyrosine does not affect the kinetics of electron transfer from heme3 to P, recent time-resolved Laue diffraction studies reported displacement of Tyr-L162 in response to the formation of the photo-oxidized P+•, implying a possible tyrosine deprotonation event. pKa values for Tyr-L162 were calculated using the corresponding crystal structures. Movement of deprotonated Tyr-L162 toward Thr-M185 was observed in P+• formation. It was associated with rearrangement of the H-bond network that proceeds to P via Thr-M185 and His-L168

Proton-Binding Sites of Acid-Sensing Ion Channel 1

Acid-sensing ion channels (ASICs) are proton-gated cation channels that exist throughout the mammalian central and peripheral nervous systems. ASIC1 is the most abundant of all the ASICs and is likely to modulate synaptic transmission. Identifying the proton-binding sites of ASCI1 is required to elucidate its pH-sensing mechanism. By using the crystal structure of ASIC1, the protonation states of each titratable site of ASIC1 were calculated by solving the Poisson-Boltzmann equation under conditions wherein the protonation states of all these sites are simultaneously in equilibrium. Four acidic-acidic residue pairs—Asp238-Asp350, Glu220-Asp408, Glu239-Asp346, and Glu80-Glu417—were found to be highly protonated. In particular, the Glu80-Glu417 pair in the inner pore was completely protonated and possessed 2 H+, implying its possible importance as a proton-binding site. The pKa of Glu239, which forms a pair with a possible pH-sensing site Asp346, differs among each homo-trimer subunit due to the different H-bond pattern of Thr237 in the different protein conformations of the subunits. His74 possessed a pKa of ≈6–7. Conservation of His74 in the proton-sensitive ASIC3 that lacks a residue corresponding to Asp346 may suggest its possible pH-sensing role in proton-sensitive ASICs

Proton transfer pathway in anion channelrhodopsin-1

Anion channelrhodopsin from Guillardia theta (GtACR1) has Asp234 (3.2 angstrom) and Glu68 (5.3 angstrom) near the protonated Schiff base. Here, we investigate mutant GtACR1s (e.g., E68Q/D234N) expressed in HEK293 cells. The influence of the acidic residues on the absorption wavelengths was also analyzed using a quantum mechanical/molecular mechanical approach. The calculated protonation pattern indicates that Asp234 is deprotonated and Glu68 is protonated in the original crystal structures. The D234E mutation and the E68Q/D234N mutation shorten and lengthen the measured and calculated absorption wavelengths, respectively, which suggests that Asp234 is deprotonated in the wild-type GtACR1. Molecular dynamics simulations show that upon mutation of deprotonated Asp234 to asparagine, deprotonated Glu68 reorients toward the Schiff base and the calculated absorption wavelength remains unchanged. The formation of the proton transfer pathway via Asp234 toward Glu68 and the disconnection of the anion conducting channel are likely a basis of the gating mechanism

Energetics of proton release on the first oxidation step in the water-oxidizing enzyme

In photosystem II (PSII), the Mn4CaO5 cluster catalyses the water splitting reaction. The crystal structure of PSII shows the presence of a hydrogen-bonded water molecule directly linked to O4. Here we show the detailed properties of the H-bonds associated with the Mn4CaO5 cluster using a quantum mechanical/molecular mechanical approach. When O4 is taken as a μ-hydroxo bridge acting as a hydrogen-bond donor to water539 (W539), the S0 redox state best describes the unusually short O4–OW539 distance (2.5 Å) seen in the crystal structure. We find that in S1, O4 easily releases the proton into a chain of eight strongly hydrogen-bonded water molecules. The corresponding hydrogen-bond network is absent for O5 in S1. The present study suggests that the O4-water chain could facilitate the initial deprotonation event in PSII. This unexpected insight is likely to be of real relevance to mechanistic models for water oxidation.UTokyo Research掲載「光合成の水分解反応初期に水素イオンが放出される仕組みを解明」 URI: http://www.u-tokyo.ac.jp/ja/utokyo-research/research-news/pathway-for-initial-proton-released-from-water-oxidizing-enzyme.htmlUTokyo Research "Pathway for initial proton released from water-oxidizing enzyme" URI: http://www.u-tokyo.ac.jp/en/utokyo-research/research-news/pathway-for-initial-proton-released-from-water-oxidizing-enzyme.htm

D139N mutation of PsbP enhances the oxygen-evolving activity of photosystem II through stabilized binding of a chloride ion

植物の光合成初期過程の酸素発生活性を向上させるアミノ酸変異を発見 --光合成・人工光合成の光エネルギー変換効率の向上へ期待--. 京都大学プレスリリース. 2022-08-18.Photosystem II (PSII) is a multi-subunit membrane protein complex that catalyzes light-driven oxidation of water to molecular oxygen. The chloride ion (Cl−) has long been known as an essential cofactor for oxygen evolution by PSII, and two Cl− ions (Cl-1 and Cl-2) have been found to specifically bind near the Mn4CaO5 cluster within the oxygen-evolving center (OEC). However, despite intensive studies on these Cl− ions, little is known about the function of Cl-2, the Cl− ion that is associated with the backbone nitrogens of D1-Asn338, D1-Phe339, and CP43-Glu354. In green plant PSII, the membrane extrinsic subunits—PsbP and PsbQ—are responsible for Cl− retention within the OEC. The Loop 4 region of PsbP, consisting of highly conserved residues Thr135–Gly142, is inserted close to Cl-2, but its importance has not been examined to date. Here, we investigated the importance of PsbP-Loop 4 using spinach PSII membranes reconstituted with spinach PsbP proteins harboring mutations in this region. Mutations in PsbP-Loop 4 had remarkable effects on the rate of oxygen evolution by PSII. Moreover, we found that a specific mutation, PsbP-D139N, significantly enhanced the oxygen-evolving activity in the absence of PsbQ, but not significantly in its presence. The D139N mutation increased the Cl− retention ability of PsbP and induced a unique structural change in the OEC, as indicated by light-induced Fourier transform infrared (FTIR) difference spectroscopy and theoretical calculations. Our findings provide insight into the functional significance of Cl-2 in the water-oxidizing reaction of PSII

Vectorial Proton Transport Mechanism of RxR, a Phylogenetically Distinct and Thermally Stable Microbial Rhodopsin

Rubrobacter xylanophilus rhodopsin (RxR) is a phylogenetically distinct and thermally stable seven-transmembrane protein that functions as a light-driven proton (H+) pump with the chromophore retinal. To characterize its vectorial proton transport mechanism, mutational and theoretical investigations were performed for carboxylates in the transmembrane region of RxR and the sequential proton transport steps were revealed as follows: (i) a proton of the retinylidene Schiff base (Lys209) is transferred to the counterion Asp74 upon formation of the blue-shifted M-intermediate in collaboration with Asp205, and simultaneously, a respective proton is released from the proton releasing group (Glu187/Glu197) to the extracellular side, (ii) a proton of Asp85 is transferred to the Schiff base during M-decay, (iii) a proton is taken up from the intracellular side to Asp85 during decay of the red-shifted O-intermediate. This ion transport mechanism of RxR provides valuable information to understand other ion transporters since carboxylates are generally essential for their functions

Characterization of tryptophan oxidation affecting D1 degradation by FtsH in the photosystem II quality control of chloroplasts

Photosynthesis is one of the most important reactions for sustaining our environment. Photosystem II (PSII) is the initial site of photosynthetic electron transfer by water oxidation. Light in excess, however, causes the simultaneous production of reactive oxygen species (ROS), leading to photo-oxidative damage in PSII. To maintain photosynthetic activity, the PSII reaction center protein D1, which is the primary target of unavoidable photo-oxidative damage, is efficiently degraded by FtsH protease. In PSII subunits, photo-oxidative modifications of several amino acids such as Trp have been indeed documented, whereas the linkage between such modifications and D1 degradation remains elusive. Here, we show that an oxidative post-translational modification of Trp residue at the N-terminal tail of D1 is correlated with D1 degradation by FtsH during high-light stress. We revealed that Arabidopsis mutant lacking FtsH2 had increased levels of oxidative Trp residues in D1, among which an N-terminal Trp-14 was distinctively localized in the stromal side. Further characterization of Trp-14 using chloroplast transformation in Chlamydomonas indicated that substitution of D1 Trp-14 to Phe, mimicking Trp oxidation enhanced FtsH-mediated D1 degradation under high light, although the substitution did not affect protein stability and PSII activity. Molecular dynamics simulation of PSII implies that both Trp-14 oxidation and Phe substitution cause fluctuation of D1 N-terminal tail. Furthermore, Trp-14 to Phe modification appeared to have an additive effect in the interaction between FtsH and PSII core in vivo. Together, our results suggest that the Trp oxidation at its N-terminus of D1 may be one of the key oxidations in the PSII repair, leading to processive degradation by FtsH

Structural basis for high selectivity of a rice silicon channel Lsi1

Silicon (Si), the most abundant mineral element in the earth’s crust, is taken up by plant roots in the form of silicic acid through Low silicon rice 1 (Lsi1). Lsi1 belongs to the Nodulin 26-like intrinsic protein subfamily in aquaporin and shows high selectivity for silicic acid. To uncover the structural basis for this high selectivity, here we show the crystal structure of the rice Lsi1 at a resolution of 1.8 Å. The structure reveals transmembrane helical orientations different from other aquaporins, characterized by a unique, widely opened, and hydrophilic selectivity filter (SF) composed of five residues. Our structural, functional, and theoretical investigations provide a solid structural basis for the Si uptake mechanism in plants, which will contribute to secure and sustainable rice production by manipulating Lsi1 selectivity for different metalloids

Origin of the pKa shift of the catalytic lysine in acetoacetate decarboxylase.

The pKa value of Lys115, the catalytic residue in acetoacetate decarboxylate, was calculated using atomic coordinates of the X-ray crystal structure with consideration of the protonation states of all titratable sites in the protein. The calculated pKa value of Lys115 (pKa(Lys115)) was unusually low (approximately 6) in agreement with the experimentally measured value. Although charged residues impact pKa(Lys115) considerably in the native protein, the significant pKa(Lys115) downshift in the protein with respect to aqueous solution was mainly due to loss of the solvation energy in the catalytic active site relative to bulk water