Molecular Plant Letter to the Editor
Crystal Structure Analysis of Extrinsic PsbP Protein of Photosystem II Reveals a ManganeseInduced Conformational Change Dear Editor, The Mn cluster (Mn4CaO5) on the thylakoid luminal side of photosystem II (PSII) catalyzes the photosynthetic oxygen-evolving reaction, an essential process for life on Earth. In higher plants and green algae, the Mn cluster is surrounded by the membraneextrinsic proteins PsbO (33 kDa), PsbP (23 kDa), and PsbQ (17 kDa) (Murata and Miyao, 1985). They shield the Mn cluster, protecting and stabilizing it from attack by exogenous reductants. PsbP plays an indispensable role in PSII function, and is necessary for the retention of Ca2+ and Cl– (Ghanotakis et al., 1984), the assembly of PSII complex (Yi et al., 2007), and the maintenance of normal thylakoid architecture (Yi et al., 2009). PsbP purified from Spinacia oleracea contained Mn ions, suggesting its involvement in providing Mn to PSII during the process of Mn cluster assembly (Bondarava et al., 2005). In addition, two crystal structures of PsbP were reported earlier, one from Nicotiana tabacum (tPsbP, Protein Data Bank [PDB] ID 1V2B; Ifuku et al., 2004) and the other from S. oleracea (sPsbP-Zn, PDB ID 2VU4; Kopecky et al., 2012). However, both were expressed in Escherichia coli, with the two internal segments unresolved in either structure, the long region (amino acids [aa] 90–107) and the short region (aa 134–139). Importantly, neither structure was found to bind Mn. Therefore, the complete structure, together with the binding sites and the coordination environment of the Mn ions within PsbP remained unknown. Here, we present two high-resolution crystal structures of the extrinsic PsbP proteins of PSII at 1.8 and 1.6 A˚ resolution, purified from S. oleracea and Zea mays, respectively (detailed methods described in Supplemental Materials and Methods). Our structures add crucial structural information for PsbP, with two Mn ions identified inside PsbP for the first time. We propose that one of the ions is responsible for a dramatic conformational change (with a maximum move of 20 A˚) that is revealed here within the long region (aa 90–107). The overall structure of spinach PsbP (sPsbP-Mn) exhibits a b-sandwich folding pattern, a common feature of members of the PsbP superfamily (data collection and refinement statistics are shown in Supplemental Table 1, protein sequence is shown in Supplemental Figure 1). Except for the N-terminal 11 residues, we constructed the structure of spinach PsbP in its entirety for the first time (P12 to A186) (Figure 1A). The two internal segments (aa 90–107 and aa 134–139), unresolved in the previous structures, were unambiguously defined in our structure based on their electron density maps (Supplemental Figure 2), with both segments exhibiting a loop conformation. Two strong electron densities were identified in the Fo-Fc and 2Fo-Fc maps of the sPsbP-Mn structure, implying the presence 664
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of two metal ions. Because spinach PsbP was incubated with 1 mM MnCl2 solution prior to crystallization, we deduced that the electron densities belonged to two Mn ions. We confirmed this assumption using the crystal X-ray fluorescence scan and the anomalous difference Fourier maps (Figure 1B and 1C). The two Mn ions were defined as Mn1 and Mn2 here. In one asymmetric unit, Mn1 is bound in a positively charged pocket (Supplemental Figure 3) and coordinated by a Cl ion and the side chains of H144 and D165, with the distances of 2.00, 2.00, and 1.86 A˚ (Figure 1B). Mn2 is coordinated by the side chain of D98 and one water molecule, with the coordinate distances of 1.96 and 2.13 A˚, respectively (Figure 1C). Notably, the Mn2coordinating residue D98 is located in the negatively charged loop at aa 90–107 (Figure 1D). In addition, we solved the crystal structure of maize PsbP (mPsbP) at 1.6 A˚ resolution (data collection and refinement statistics shown in Supplemental Table 1, protein sequence shown in Supplemental Figure 1). There are two PsbP molecules in one asymmetric unit, which exhibit similar structures, with a root-mean-square deviation value of 0.39 A˚ for their Ca atoms (Figure 1E). No metal ions were found in this structure. The residues from N15 to A186 of the mPsbP structure were modeled, except for the region from S99 to F103, as it was not visible on the electron density map. To perform more detailed structural analysis, we superposed all four PsbP structures, including the two structures reported previously. We found that the structures are all folded in an identical b-sandwich pattern. However, a number of structural features exhibit varying degrees of divergence among the four structures. Differences are found in regions of the N terminus, as well as specific loops, i.e. loop90–107, loop120–125, loop134–140, and loop168–173 (the corresponding regions are shown in Figure 1F). The structure of sPsbP-Mn possesses a longer N terminus, starting at the residue P12, whereas the N termini of the remaining structures start from the 15th residue. It was reported earlier that the N-terminal region of PsbP was essential for the functional reconstitution of the PSII complex (Tomita et al., 2009), and that it directly interacted with PsbE (Cyt b559 a subunit) within PSII (Ido et al., 2012). In the sPsbP-Mn structure, the N terminus exhibits a loop conformation and stretches out from its core domain. Among four structures, only our sPsbP-Mn structure and the sPsbP-Zn structure previously resolved (PDB ID 2VU4) were
Published by the Molecular Plant Shanghai Editorial Office in association with Cell Press, an imprint of Elsevier Inc., on behalf of CSPB and IPPE, SIBS, CAS.
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Figure 1. Crystal Structures of Extrinsic PsbP Protein of Photosystem II Purified from Spinacia oleracea and Zea mays. (A) Cartoon ribbon diagram of Mn-bound spinach PsbP, with Mn (Mn1 and Mn2) and Cl ions shown as spheres. Residues H144, D165, and D98, which coordinate with two Mn ions, are shown as sticks. Two segments (loop90–107 and loop134–139) unresolved in earlier structures are shown in pink and white. (B) Coordination of Mn1 with H144, D165, and a Cl ion (green sphere). (C) Coordination of Mn2 with D98 and a water molecule (red sphere). In (B) and (C), the anomalous difference Fourier maps (blue, contour at 10s) are shown for Mn1 and Mn2. (D) sPsbP-Mn (hot pink) is superimposed with the structure of sPsbP-Zn (lime). Zn ion locates at the same site as Mn1. (E) Cartoon ribbon diagram of maize PsbP. Two molecules in one asymmetry unit of crystal structure were superposed. The dotted line represents the unbuilt region of residues 99–103. (F) Superposition of four structures of PsbP proteins from different plants. mPsbP, PsbP from Zea mays; sPsbP-Mn, Mn-bound PsbP from Spinacia oleracea; sPsbP-Zn, 2VU4 from Spinacia oleracea; tPsbP, 1V2B from Nicotiana tabacum. Loops with different conformations are indicated. (G) Locally enlarged view of two different conformations of loop90–107. Upon binding of Mn ions, loop90–107 swings around by approximately 90 , with a distance between two D98 residues (shown as sticks) of approximately 20 A˚.
found to bind metal ions. However, their metal-binding properties appear to differ. sPsbP-Mn binds two Mn ions, whereas sPsbPZn contains only one single Zn ion, which is located at the same site of Mn1 (Figure 1D). The Zn ion exhibits a similar binding mode with Mn1, but the coordination distances are longer than those of Mn1. It is noteworthy that Zn ions interfere with PsbP function, as they caused the dissociation of PsbP and PsbQ from PSII core, and simultaneously inhibited the oxygen-evolving activity of PSII in higher plants (Rashid et al., 1994). In contrast, under physiological conditions, Mn ions are essential for proper functioning of the Mn cluster in PSII complex. Moreover, they were co-purified with the PsbP protein from spinach (Bondarava et al., 2005). Together, these observations are in agreement with our experimental results, which showed that naturally purified PsbP protein was unstable and tended to precipitate upon exposure to even low concentrations of Zn ions (1 mM). In contrast, PsbP remained in solution following addition of Mn ions (1 mM). In conclusion,
these results suggest that our Mn-bound structure represents a more natural PsbP conformation under physiological conditions than the Zn-bound structure. To understand the precise role of Mn ions, we performed a more detailed structural analysis of the interactions between Mn and PsbP. In our sPsbP-Mn and mPsbP structures, detailed comparison reveals that upon Mn binding, loop90–107 undergoes a significant conformational change (Figure 1F and 1G). In mPsbP, a metal-free structure, loop90–107 is extending to the outer part of the protein. In the Mn-bound structure (sPsbP-Mn), however, one Mn ion (Mn2) coordinates to the side chain of D98, which induces loop90–107 to swing around by approximately 90 . As a result, loop90–107 becomes positioned closer to the b-sandwich core of PsbP when compared with its position in the metal-free form. The distance between two D98 residues in the Mn-bound and metal-free structures is approximately 20 A˚. In the Mn1 site, in contrast, no significant difference between the two Molecular Plant 8, 664–666, April 2015 ª The Author 2015.
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Molecular Plant structures is observed (Supplemental Figure 4). We propose that the observed conformational change in the loop90–107 region is induced by binding of Mn2. Interestingly, Bondarava et al. (2007) reported a stoichiometry of 1:1 between the recombinant PsbP protein and Mn ions, yet in the same study they proposed that an unstable functional intermediate was formed, in which one PsbP contained two Mn ions. In our sPsbP-Mn structure, two Mn ions were clearly identified from the crystal X-ray fluorescence scan and the anomalous difference Fourier maps, demonstrating that the PsbP protein has two binding sites of Mn ions. Based on the coordination modes of Mn ions, we propose that Mn1 represents the primary binding site of high affinity, whereas Mn2 is a secondary binding site with relatively low affinity. The Mn ion at the Mn2 site is less stable than that at the Mn1 site, and was possibly lost during the dialysis step of the purification procedure of recombinant PsbP (Bondarava et al., 2007). Furthermore, it was proposed that PsbP would provide Mn ions for the Mn cluster on the thylakoid luminal side of PSII (Bondarava et al., 2007). As a stable coordination site, Mn1 may not be able to execute this function. In contrast, the Mn2 site, with its low affinity to Mn ions, would be a suitable candidate to donate Mn ions. Combined with those previous biochemical results, we propose here that Mnbound and metal-free PsbP structures represent two different conformations during the PSII damage–repair cycle. Thus, PsbP will function as a carrier for Mn, facilitating the access of Mn ions to the oxygen-evolving Mn cluster of PSII, namely by way of a major conformational change within the flexible loop90–107. Furthermore, based on the evidence presented here, and in combination with insights from earlier studies, we propose a novel docking model for the location of PsbP in higher plant PSII (a detailed discussion of our model is provided in the Supplemental Discussion).
DATA DEPOSITION The atomic coordinates and structure factors have been deposited in the Protein Data Bank (http://www.pdb.org) under accession codes 4RTH and 4RTI.
SUPPLEMENTAL INFORMATION Supplemental Information is available at Molecular Plant Online.
FUNDING This research was financially supported by grants from the National Natural Science Foundation of China (Grant No. 31100534, 31021062 and 31170703), the Ministry of Science and Technology (973 Programs, Grant No. 2011CBA00902), and the Strategic Priority Research program of the Chinese Academy of Sciences (XDB08020302).
ACKNOWLEDGMENTS We thank Alberta Pinnola for technical assistance on the maize cultivation and PSII-BBY extraction. We thank Zhenfeng Liu, Minrui Fan, and Jiping Zhang for helpful discussion. We are grateful to Yi Han and Shengquan Liu at the Institute of Biophysics, CAS, the staff of beamline BL17U at the Shanghai Synchrotron Radiation Facility, and the staff of beamlines BL1A, BL17A, and NW12A at the Photon Factory, KEK (Tsukuba, Japan) for technical support. No conflict of interest declared.
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Molecular Plant 8, 664–666, April 2015 ª The Author 2015.
Letter to the Editor Received: November 16, 2014 Revised: December 30, 2014 Accepted: January 5, 2015 Published: January 9, 2015
Peng Cao1,3, Yuan Xie1,2,3, Mei Li1,*, Xiaowei Pan1, Hongmei Zhang1, Xuelin Zhao1, Xiaodong Su1, Tao Cheng1,2 and Wenrui Chang1,* 1
National Laboratory of Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, 15th Datun Road, Chaoyang District, Beijing 100101, China 2 University of Chinese Academy of Sciences, 19A Yuquan Road, Beijing 100049, China 3 These authors contributed equally to this article. *Correspondence: Wenrui Chang ([email protected]), Mei Li ([email protected]) http://dx.doi.org/10.1016/j.molp.2015.01.002 This is an open access article under the CC BY license (http:// creativecommons.org/licenses/by/4.0/).
REFERENCES Bondarava, N., Beyer, P., and Krieger-Liszkay, A. (2005). Function of the 23 kDa extrinsic protein of photosystem II as a manganese binding protein and its role in photoactivation. Biochim. Biophys. Acta 1708:63–70. Bondarava, N., Un, S., and Krieger-Liszkay, A. (2007). Manganese binding to the 23 kDa extrinsic protein of Photosystem II. Biochim. Biophys. Acta 1767:583–588. Ghanotakis, D.F., Topper, J.N., Babcock, G.T., and Yocum, C.F. (1984). Water-soluble 17 and 23 kDa polypeptides restore oxygen evolution activity by creating a high-affinity binding site for Ca2 on the oxidizing side of photosystem II. FEBS Lett. 170:169–173. Ido, K., Kakiuchi, S., Uno, C., Nishimura, T., Fukao, Y., Noguchi, T., Sato, F., and Ifuku, K. (2012). The conserved His-144 in the PsbP protein is important for the interaction between the PsbP N-terminus and the Cyt b(559) subunit of photosystem II. J. Biol. Chem. 287:26377–26387. Ifuku, K., Nakatsu, T., Kato, H., and Sato, F. (2004). Crystal structure of the PsbP protein of photosystem II from Nicotiana tabacum. EMBO Rep. 5:362–367. Kopecky, V., Jr., Kohoutova, J., Lapkouski, M., Hofbauerova, K., Sovova, Z., Ettrichova, O., Gonzalez-Perez, S., Dulebo, A., Kaftan, D., Smatanova, I.K., et al. (2012). Raman spectroscopy adds complementary detail to the high-resolution x-ray crystal structure of photosynthetic PsbP from Spinacia oleracea. PLoS One 7:e46694. Murata, N., and Miyao, M. (1985). Extrinsic membrane proteins in the photosynthetic oxygen-evolving complex. Trends Biochem. Sci. 10:122–124. Rashid, A., Camm, E.L., and Ekramoddoullah, A.K. (1994). Molecular mechanism of action of Pb2+ and Zn2+ on water oxidizing complex of photosystem II. FEBS Lett. 350:296–298. Tomita, M., Ifuku, K., Sato, F., and Noguchi, T. (2009). FTIR evidence that the PsbP extrinsic protein induces protein conformational changes around the oxygen-evolving Mn cluster in photosystem II. Biochemistry 48:6318–6325. Yi, X., Hargett, S.R., Liu, H., Frankel, L.K., and Bricker, T.M. (2007). The PsbP protein is required for photosystem II complex assembly/ stability and photoautotrophy in Arabidopsis thaliana. J. Biol. Chem. 282:24833–24841. Yi, X., Hargett, S.R., Frankel, L.K., and Bricker, T.M. (2009). The PsbP protein, but not the PsbQ protein, is required for normal thylakoid architecture in Arabidopsis thaliana. FEBS Lett. 583 (12):2142–2147.
Crystal Structure Analysis of Extrinsic PsbP Protein of Photosystem II Reveals a ManganeseInduced Conformational Change Dear Editor, The Mn cluster (Mn4CaO5) on the thylakoid luminal side of photosystem II (PSII) catalyzes the photosynthetic oxygen-evolving reaction, an essential process for life on Earth. In higher plants and green algae, the Mn cluster is surrounded by the membraneextrinsic proteins PsbO (33 kDa), PsbP (23 kDa), and PsbQ (17 kDa) (Murata and Miyao, 1985). They shield the Mn cluster, protecting and stabilizing it from attack by exogenous reductants. PsbP plays an indispensable role in PSII function, and is necessary for the retention of Ca2+ and Cl– (Ghanotakis et al., 1984), the assembly of PSII complex (Yi et al., 2007), and the maintenance of normal thylakoid architecture (Yi et al., 2009). PsbP purified from Spinacia oleracea contained Mn ions, suggesting its involvement in providing Mn to PSII during the process of Mn cluster assembly (Bondarava et al., 2005). In addition, two crystal structures of PsbP were reported earlier, one from Nicotiana tabacum (tPsbP, Protein Data Bank [PDB] ID 1V2B; Ifuku et al., 2004) and the other from S. oleracea (sPsbP-Zn, PDB ID 2VU4; Kopecky et al., 2012). However, both were expressed in Escherichia coli, with the two internal segments unresolved in either structure, the long region (amino acids [aa] 90–107) and the short region (aa 134–139). Importantly, neither structure was found to bind Mn. Therefore, the complete structure, together with the binding sites and the coordination environment of the Mn ions within PsbP remained unknown. Here, we present two high-resolution crystal structures of the extrinsic PsbP proteins of PSII at 1.8 and 1.6 A˚ resolution, purified from S. oleracea and Zea mays, respectively (detailed methods described in Supplemental Materials and Methods). Our structures add crucial structural information for PsbP, with two Mn ions identified inside PsbP for the first time. We propose that one of the ions is responsible for a dramatic conformational change (with a maximum move of 20 A˚) that is revealed here within the long region (aa 90–107). The overall structure of spinach PsbP (sPsbP-Mn) exhibits a b-sandwich folding pattern, a common feature of members of the PsbP superfamily (data collection and refinement statistics are shown in Supplemental Table 1, protein sequence is shown in Supplemental Figure 1). Except for the N-terminal 11 residues, we constructed the structure of spinach PsbP in its entirety for the first time (P12 to A186) (Figure 1A). The two internal segments (aa 90–107 and aa 134–139), unresolved in the previous structures, were unambiguously defined in our structure based on their electron density maps (Supplemental Figure 2), with both segments exhibiting a loop conformation. Two strong electron densities were identified in the Fo-Fc and 2Fo-Fc maps of the sPsbP-Mn structure, implying the presence 664
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of two metal ions. Because spinach PsbP was incubated with 1 mM MnCl2 solution prior to crystallization, we deduced that the electron densities belonged to two Mn ions. We confirmed this assumption using the crystal X-ray fluorescence scan and the anomalous difference Fourier maps (Figure 1B and 1C). The two Mn ions were defined as Mn1 and Mn2 here. In one asymmetric unit, Mn1 is bound in a positively charged pocket (Supplemental Figure 3) and coordinated by a Cl ion and the side chains of H144 and D165, with the distances of 2.00, 2.00, and 1.86 A˚ (Figure 1B). Mn2 is coordinated by the side chain of D98 and one water molecule, with the coordinate distances of 1.96 and 2.13 A˚, respectively (Figure 1C). Notably, the Mn2coordinating residue D98 is located in the negatively charged loop at aa 90–107 (Figure 1D). In addition, we solved the crystal structure of maize PsbP (mPsbP) at 1.6 A˚ resolution (data collection and refinement statistics shown in Supplemental Table 1, protein sequence shown in Supplemental Figure 1). There are two PsbP molecules in one asymmetric unit, which exhibit similar structures, with a root-mean-square deviation value of 0.39 A˚ for their Ca atoms (Figure 1E). No metal ions were found in this structure. The residues from N15 to A186 of the mPsbP structure were modeled, except for the region from S99 to F103, as it was not visible on the electron density map. To perform more detailed structural analysis, we superposed all four PsbP structures, including the two structures reported previously. We found that the structures are all folded in an identical b-sandwich pattern. However, a number of structural features exhibit varying degrees of divergence among the four structures. Differences are found in regions of the N terminus, as well as specific loops, i.e. loop90–107, loop120–125, loop134–140, and loop168–173 (the corresponding regions are shown in Figure 1F). The structure of sPsbP-Mn possesses a longer N terminus, starting at the residue P12, whereas the N termini of the remaining structures start from the 15th residue. It was reported earlier that the N-terminal region of PsbP was essential for the functional reconstitution of the PSII complex (Tomita et al., 2009), and that it directly interacted with PsbE (Cyt b559 a subunit) within PSII (Ido et al., 2012). In the sPsbP-Mn structure, the N terminus exhibits a loop conformation and stretches out from its core domain. Among four structures, only our sPsbP-Mn structure and the sPsbP-Zn structure previously resolved (PDB ID 2VU4) were
Published by the Molecular Plant Shanghai Editorial Office in association with Cell Press, an imprint of Elsevier Inc., on behalf of CSPB and IPPE, SIBS, CAS.
Molecular Plant
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Figure 1. Crystal Structures of Extrinsic PsbP Protein of Photosystem II Purified from Spinacia oleracea and Zea mays. (A) Cartoon ribbon diagram of Mn-bound spinach PsbP, with Mn (Mn1 and Mn2) and Cl ions shown as spheres. Residues H144, D165, and D98, which coordinate with two Mn ions, are shown as sticks. Two segments (loop90–107 and loop134–139) unresolved in earlier structures are shown in pink and white. (B) Coordination of Mn1 with H144, D165, and a Cl ion (green sphere). (C) Coordination of Mn2 with D98 and a water molecule (red sphere). In (B) and (C), the anomalous difference Fourier maps (blue, contour at 10s) are shown for Mn1 and Mn2. (D) sPsbP-Mn (hot pink) is superimposed with the structure of sPsbP-Zn (lime). Zn ion locates at the same site as Mn1. (E) Cartoon ribbon diagram of maize PsbP. Two molecules in one asymmetry unit of crystal structure were superposed. The dotted line represents the unbuilt region of residues 99–103. (F) Superposition of four structures of PsbP proteins from different plants. mPsbP, PsbP from Zea mays; sPsbP-Mn, Mn-bound PsbP from Spinacia oleracea; sPsbP-Zn, 2VU4 from Spinacia oleracea; tPsbP, 1V2B from Nicotiana tabacum. Loops with different conformations are indicated. (G) Locally enlarged view of two different conformations of loop90–107. Upon binding of Mn ions, loop90–107 swings around by approximately 90 , with a distance between two D98 residues (shown as sticks) of approximately 20 A˚.
found to bind metal ions. However, their metal-binding properties appear to differ. sPsbP-Mn binds two Mn ions, whereas sPsbPZn contains only one single Zn ion, which is located at the same site of Mn1 (Figure 1D). The Zn ion exhibits a similar binding mode with Mn1, but the coordination distances are longer than those of Mn1. It is noteworthy that Zn ions interfere with PsbP function, as they caused the dissociation of PsbP and PsbQ from PSII core, and simultaneously inhibited the oxygen-evolving activity of PSII in higher plants (Rashid et al., 1994). In contrast, under physiological conditions, Mn ions are essential for proper functioning of the Mn cluster in PSII complex. Moreover, they were co-purified with the PsbP protein from spinach (Bondarava et al., 2005). Together, these observations are in agreement with our experimental results, which showed that naturally purified PsbP protein was unstable and tended to precipitate upon exposure to even low concentrations of Zn ions (1 mM). In contrast, PsbP remained in solution following addition of Mn ions (1 mM). In conclusion,
these results suggest that our Mn-bound structure represents a more natural PsbP conformation under physiological conditions than the Zn-bound structure. To understand the precise role of Mn ions, we performed a more detailed structural analysis of the interactions between Mn and PsbP. In our sPsbP-Mn and mPsbP structures, detailed comparison reveals that upon Mn binding, loop90–107 undergoes a significant conformational change (Figure 1F and 1G). In mPsbP, a metal-free structure, loop90–107 is extending to the outer part of the protein. In the Mn-bound structure (sPsbP-Mn), however, one Mn ion (Mn2) coordinates to the side chain of D98, which induces loop90–107 to swing around by approximately 90 . As a result, loop90–107 becomes positioned closer to the b-sandwich core of PsbP when compared with its position in the metal-free form. The distance between two D98 residues in the Mn-bound and metal-free structures is approximately 20 A˚. In the Mn1 site, in contrast, no significant difference between the two Molecular Plant 8, 664–666, April 2015 ª The Author 2015.
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Molecular Plant structures is observed (Supplemental Figure 4). We propose that the observed conformational change in the loop90–107 region is induced by binding of Mn2. Interestingly, Bondarava et al. (2007) reported a stoichiometry of 1:1 between the recombinant PsbP protein and Mn ions, yet in the same study they proposed that an unstable functional intermediate was formed, in which one PsbP contained two Mn ions. In our sPsbP-Mn structure, two Mn ions were clearly identified from the crystal X-ray fluorescence scan and the anomalous difference Fourier maps, demonstrating that the PsbP protein has two binding sites of Mn ions. Based on the coordination modes of Mn ions, we propose that Mn1 represents the primary binding site of high affinity, whereas Mn2 is a secondary binding site with relatively low affinity. The Mn ion at the Mn2 site is less stable than that at the Mn1 site, and was possibly lost during the dialysis step of the purification procedure of recombinant PsbP (Bondarava et al., 2007). Furthermore, it was proposed that PsbP would provide Mn ions for the Mn cluster on the thylakoid luminal side of PSII (Bondarava et al., 2007). As a stable coordination site, Mn1 may not be able to execute this function. In contrast, the Mn2 site, with its low affinity to Mn ions, would be a suitable candidate to donate Mn ions. Combined with those previous biochemical results, we propose here that Mnbound and metal-free PsbP structures represent two different conformations during the PSII damage–repair cycle. Thus, PsbP will function as a carrier for Mn, facilitating the access of Mn ions to the oxygen-evolving Mn cluster of PSII, namely by way of a major conformational change within the flexible loop90–107. Furthermore, based on the evidence presented here, and in combination with insights from earlier studies, we propose a novel docking model for the location of PsbP in higher plant PSII (a detailed discussion of our model is provided in the Supplemental Discussion).
DATA DEPOSITION The atomic coordinates and structure factors have been deposited in the Protein Data Bank (http://www.pdb.org) under accession codes 4RTH and 4RTI.
SUPPLEMENTAL INFORMATION Supplemental Information is available at Molecular Plant Online.
FUNDING This research was financially supported by grants from the National Natural Science Foundation of China (Grant No. 31100534, 31021062 and 31170703), the Ministry of Science and Technology (973 Programs, Grant No. 2011CBA00902), and the Strategic Priority Research program of the Chinese Academy of Sciences (XDB08020302).
ACKNOWLEDGMENTS We thank Alberta Pinnola for technical assistance on the maize cultivation and PSII-BBY extraction. We thank Zhenfeng Liu, Minrui Fan, and Jiping Zhang for helpful discussion. We are grateful to Yi Han and Shengquan Liu at the Institute of Biophysics, CAS, the staff of beamline BL17U at the Shanghai Synchrotron Radiation Facility, and the staff of beamlines BL1A, BL17A, and NW12A at the Photon Factory, KEK (Tsukuba, Japan) for technical support. No conflict of interest declared.
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Molecular Plant 8, 664–666, April 2015 ª The Author 2015.
Letter to the Editor Received: November 16, 2014 Revised: December 30, 2014 Accepted: January 5, 2015 Published: January 9, 2015
Peng Cao1,3, Yuan Xie1,2,3, Mei Li1,*, Xiaowei Pan1, Hongmei Zhang1, Xuelin Zhao1, Xiaodong Su1, Tao Cheng1,2 and Wenrui Chang1,* 1
National Laboratory of Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, 15th Datun Road, Chaoyang District, Beijing 100101, China 2 University of Chinese Academy of Sciences, 19A Yuquan Road, Beijing 100049, China 3 These authors contributed equally to this article. *Correspondence: Wenrui Chang ([email protected]), Mei Li ([email protected]) http://dx.doi.org/10.1016/j.molp.2015.01.002 This is an open access article under the CC BY license (http:// creativecommons.org/licenses/by/4.0/).
REFERENCES Bondarava, N., Beyer, P., and Krieger-Liszkay, A. (2005). Function of the 23 kDa extrinsic protein of photosystem II as a manganese binding protein and its role in photoactivation. Biochim. Biophys. Acta 1708:63–70. Bondarava, N., Un, S., and Krieger-Liszkay, A. (2007). Manganese binding to the 23 kDa extrinsic protein of Photosystem II. Biochim. Biophys. Acta 1767:583–588. Ghanotakis, D.F., Topper, J.N., Babcock, G.T., and Yocum, C.F. (1984). Water-soluble 17 and 23 kDa polypeptides restore oxygen evolution activity by creating a high-affinity binding site for Ca2 on the oxidizing side of photosystem II. FEBS Lett. 170:169–173. Ido, K., Kakiuchi, S., Uno, C., Nishimura, T., Fukao, Y., Noguchi, T., Sato, F., and Ifuku, K. (2012). The conserved His-144 in the PsbP protein is important for the interaction between the PsbP N-terminus and the Cyt b(559) subunit of photosystem II. J. Biol. Chem. 287:26377–26387. Ifuku, K., Nakatsu, T., Kato, H., and Sato, F. (2004). Crystal structure of the PsbP protein of photosystem II from Nicotiana tabacum. EMBO Rep. 5:362–367. Kopecky, V., Jr., Kohoutova, J., Lapkouski, M., Hofbauerova, K., Sovova, Z., Ettrichova, O., Gonzalez-Perez, S., Dulebo, A., Kaftan, D., Smatanova, I.K., et al. (2012). Raman spectroscopy adds complementary detail to the high-resolution x-ray crystal structure of photosynthetic PsbP from Spinacia oleracea. PLoS One 7:e46694. Murata, N., and Miyao, M. (1985). Extrinsic membrane proteins in the photosynthetic oxygen-evolving complex. Trends Biochem. Sci. 10:122–124. Rashid, A., Camm, E.L., and Ekramoddoullah, A.K. (1994). Molecular mechanism of action of Pb2+ and Zn2+ on water oxidizing complex of photosystem II. FEBS Lett. 350:296–298. Tomita, M., Ifuku, K., Sato, F., and Noguchi, T. (2009). FTIR evidence that the PsbP extrinsic protein induces protein conformational changes around the oxygen-evolving Mn cluster in photosystem II. Biochemistry 48:6318–6325. Yi, X., Hargett, S.R., Liu, H., Frankel, L.K., and Bricker, T.M. (2007). The PsbP protein is required for photosystem II complex assembly/ stability and photoautotrophy in Arabidopsis thaliana. J. Biol. Chem. 282:24833–24841. Yi, X., Hargett, S.R., Frankel, L.K., and Bricker, T.M. (2009). The PsbP protein, but not the PsbQ protein, is required for normal thylakoid architecture in Arabidopsis thaliana. FEBS Lett. 583 (12):2142–2147.
