Photosystem II Complex in Vivo Is a Monomer
2009; Elsevier BV; Volume: 284; Issue: 23 Linguagem: Inglês
10.1074/jbc.m109.000372
ISSN1083-351X
AutoresTakeshi Takahashi, Natsuko Inoue‐Kashino, Shinichiro Ozawa, Yuichiro Takahashi, Yasuhiro Kashino, Kazuhiko Satoh,
Tópico(s)Photoreceptor and optogenetics research
ResumoPhotosystem II (PS II) complexes are membrane protein complexes that are composed of >20 distinct subunit proteins. Similar to many other membrane protein complexes, two PS II complexes are believed to form a homo-dimer whose molecular mass is ∼650 kDa. Contrary to this well known concept, we propose that the functional form of PS II in vivo is a monomer, based on the following observations. Deprivation of lipids caused the conversion of PS II from a monomeric form to a dimeric form. Only a monomeric PS II was detected in solubilized cyanobacterial and red algal thylakoids using blue-native polyacrylamide gel electrophoresis. Furthermore, energy transfer between PS II units, which was observed in the purified dimeric PS II, was not detected in vivo. Our proposal will lead to a re-evaluation of many crystallographic models of membrane protein complexes in terms of their oligomerization status. Photosystem II (PS II) complexes are membrane protein complexes that are composed of >20 distinct subunit proteins. Similar to many other membrane protein complexes, two PS II complexes are believed to form a homo-dimer whose molecular mass is ∼650 kDa. Contrary to this well known concept, we propose that the functional form of PS II in vivo is a monomer, based on the following observations. Deprivation of lipids caused the conversion of PS II from a monomeric form to a dimeric form. Only a monomeric PS II was detected in solubilized cyanobacterial and red algal thylakoids using blue-native polyacrylamide gel electrophoresis. Furthermore, energy transfer between PS II units, which was observed in the purified dimeric PS II, was not detected in vivo. Our proposal will lead to a re-evaluation of many crystallographic models of membrane protein complexes in terms of their oligomerization status. Photosystem II (PS II) 3The abbreviations used are:PS I and PS IIphotosystem I and IIBNblue nativeChlchlorophyllClogPthe calculated logarithm of the partition coefficient2,5-DCBQ2,5-dichloro-p-benzoquinoneDCMU3-(3,4-dichlorophenyl)-1,1-dimethyl ureaDDMn-dodecyl-β-d-maltosideLHCIIlight-harvesting Chl-binding proteinsMES2-(N-morpholino)ethanesulfonic acidLCliquid chromatographyMS/MStandem mass spectrometry. 3The abbreviations used are:PS I and PS IIphotosystem I and IIBNblue nativeChlchlorophyllClogPthe calculated logarithm of the partition coefficient2,5-DCBQ2,5-dichloro-p-benzoquinoneDCMU3-(3,4-dichlorophenyl)-1,1-dimethyl ureaDDMn-dodecyl-β-d-maltosideLHCIIlight-harvesting Chl-binding proteinsMES2-(N-morpholino)ethanesulfonic acidLCliquid chromatographyMS/MStandem mass spectrometry. complexes convert solar energy to biological redox energy. Through this reaction process, water molecules are oxidized and molecular oxygen is released as a byproduct (reviewed in Ref. 1.Renger G. Renger T. Photosynth. Res. 2008; 98: 53-80Crossref PubMed Scopus (243) Google Scholar), which is the only source of molecular oxygen upon which all aerobic organisms on earth rely. PS II core complexes are membrane protein complexes that are composed of >20 distinct subunit proteins and many functional cofactors, including chlorophylls (Chls), carotenoids, plastoquinone, and metal ions (2.Ferreira K.N. Iverson T.M. Maghlaoui K. Barber J. Iwata S. Science. 2004; 303: 1831-1838Crossref PubMed Scopus (2823) Google Scholar, 3.Kamiya N. Shen J.R. Proc. Natl. Acad. Sci. U.S.A. 2003; 100: 98-103Crossref PubMed Scopus (991) Google Scholar, 4.Kashino Y. Lauber W.M. Carroll J.A. Wang Q. Whitmarsh J. Satoh K. Pakrasi H.B. Biochemistry. 2002; 41: 8004-8012Crossref PubMed Scopus (278) Google Scholar, 5.Loll B. Kern J. Saenger W. Zouni A. Biesiadka J. Nature. 2005; 438: 1040-1044Crossref PubMed Scopus (1595) Google Scholar). Similar to many other membrane protein complexes (6.Huang D. Everly R.M. Cheng R.H. Heymann J.B. Schägger H. Sled V. Ohnishi T. Baker T.S. Cramer W.A. Biochemistry. 1994; 33: 4401-4409Crossref PubMed Scopus (104) Google Scholar, 7.Kurisu G. Zhang H. Smith J.L. Cramer W.A. Science. 2003; 302: 1009-1014Crossref PubMed Scopus (582) Google Scholar, 8.Schägger H. Cramer W.A. von Jagow G. Anal. Biochem. 1994; 217: 220-230Crossref PubMed Scopus (1033) Google Scholar, 9.Stroebel D. Choquet Y. Popot J.L. Picot D. Nature. 2003; 426: 413-418Crossref PubMed Scopus (498) Google Scholar, 10.Tsukihara T. Aoyama H. Yamashita E. Tomizaki T. Yamaguchi H. Shinzawa-Itoh K. Nakashima R. Yaono R. Yoshikawa S. Science. 1995; 269: 1069-1674Crossref PubMed Scopus (1286) Google Scholar), two PS II core complexes are believed to associate together to form a homo-dimer with a molecular mass of ∼650 kDa, as shown by crystallographic models (2.Ferreira K.N. Iverson T.M. Maghlaoui K. Barber J. Iwata S. Science. 2004; 303: 1831-1838Crossref PubMed Scopus (2823) Google Scholar, 3.Kamiya N. Shen J.R. Proc. Natl. Acad. Sci. U.S.A. 2003; 100: 98-103Crossref PubMed Scopus (991) Google Scholar, 5.Loll B. Kern J. Saenger W. Zouni A. Biesiadka J. Nature. 2005; 438: 1040-1044Crossref PubMed Scopus (1595) Google Scholar).The PS II complex turns over dynamically, although it is quite an integrated complex; our current understanding is that the PS II complex that is damaged by high light is disintegrated into a monomeric form and is further dissociated to replace a degraded D1 protein with a de novo synthesized D1 (reviewed in Refs. 11.Aro E.M. Suorsa M. Rokka A. Allahverdiyeva Y. Paakkarinen V. Saleem A. Battchikova N. Rintamaki E. J. Exp. Bot. 2005; 56: 347-356Crossref PubMed Scopus (380) Google Scholar and 12.Nixon P.J. Barker M. Boehm M. de Vries R. Komenda J. J. Exp. Bot. 2005; 56: 357-363Crossref PubMed Scopus (157) Google Scholar). After the replacement, the PS II complex is integrated into a functional form as a dimer. It is supposed that PS II subunit proteins such as PsbI (13.Dobáková M. Tichy M. Komenda J. Plant Physiol. 2007; 145: 1681-1691Crossref PubMed Scopus (65) Google Scholar) or PsbTc (14.Iwai M. Katoh H. Katayama M. Ikeuchi M. Plant Cell Physiol. 2004; 45: 1809-1816Crossref PubMed Scopus (33) Google Scholar) participate in the formation of the PS II dimer.Crystallographic models of PS II have enabled the determination of the accurate molecular architecture of PS II complexes, all of which are in a dimeric form. The most recent crystallographic model of the PS II dimer at 3.0-Å resolution revealed the presence of six detergent molecules located at the interface of the two monomers (5.Loll B. Kern J. Saenger W. Zouni A. Biesiadka J. Nature. 2005; 438: 1040-1044Crossref PubMed Scopus (1595) Google Scholar). Small structural fluctuations during the purification process might allow the invasion of those detergent molecules. However, it is also probable that the PS II complexes exist in the form of a monomer in vivo and the two distinct monomers become a dimer during the purification step incorporating detergents between their interfaces. This idea led us to investigate the actual form of PS II in vivo. Contrary to the above well known dimeric model of a functional PS II core complex, here we show that the PS II core complex functions and exists in a monomeric form in vivo.DISCUSSIONIn this work, we showed that PS II functions mostly in a monomeric form in vivo, contrary to the widely accepted concept that the functional form of PS II complexes is the dimer. This conventional concept is consistent with and stands on the crystallographic models of PS II reported so far (2.Ferreira K.N. Iverson T.M. Maghlaoui K. Barber J. Iwata S. Science. 2004; 303: 1831-1838Crossref PubMed Scopus (2823) Google Scholar, 3.Kamiya N. Shen J.R. Proc. Natl. Acad. Sci. U.S.A. 2003; 100: 98-103Crossref PubMed Scopus (991) Google Scholar, 5.Loll B. Kern J. Saenger W. Zouni A. Biesiadka J. Nature. 2005; 438: 1040-1044Crossref PubMed Scopus (1595) Google Scholar, 40.Kuhl H. Kruip J. Seidler A. Krieger-Liszkay A. Bunker M. Bald D. Scheidig A.J. Rogner M. J. Biol. Chem. 2000; 275: 20652-20659Abstract Full Text Full Text PDF PubMed Scopus (88) Google Scholar, 41.Zouni A. Witt H.T. Kern J. Fromme P. Krauss N. Saenger W. Orth P. Nature. 2001; 409: 739-743Crossref PubMed Scopus (1754) Google Scholar). However, it is natural that the crystallographic models of PS II are dimers, because all of the PS II crystals were made using purified PS II dimers. One prerequisite for obtaining crystals of membrane protein complexes is to remove lipids (26.Kashino Y. J. Chromatogr. B. 2003; 797: 191-216Crossref PubMed Scopus (68) Google Scholar). As we have shown in this work (), the extensive removal of lipids, on purpose or unintentionally, during the purification steps for the crystallization work could result in the high yield of PS II dimer. The side surface opposing the interface between the two monomers in dimeric PS II might be specific to the formation of dimer, because the loss of PsbTc that locates at such an interface results in a decrease in the ratio of dimeric PS II (14.Iwai M. Katoh H. Katayama M. Ikeuchi M. Plant Cell Physiol. 2004; 45: 1809-1816Crossref PubMed Scopus (33) Google Scholar).To our knowledge, no direct evidence on the in vivo form of PS II complexes has been reported. The model of the PS II dimer is primarily based on freeze-fracture electron microscopic studies and crystallographic models. The freeze-fracture studies revealed the presence of pairs of 10 nm particles associated with phycobilisomes, which were suggested to be the dimeric forms of PS II (55.Giddings T.H. Wasmann C. Staehelin L.A. Plant Physiol. 1983; 71: 409-419Crossref PubMed Google Scholar, 56.Giddings Jr., T.H. Staehelin L.A. Biochim. Biophys. Acta. 1979; 546: 373-382Crossref PubMed Scopus (30) Google Scholar, 57.Mörschel E. Schatz G.H. Planta. 1987; 172: 145-154Crossref PubMed Scopus (62) Google Scholar). PS II monomers could associate together to form a dimer in some physiological statuses. Actually, Wollman reported that 10 nm particles observed on the exoplasmic fracture face are attributable to PS II and are apparently able to aggregate once the phycobilisomes are detached in Cyanidium caldarium (58.Wollman F.A. Plant Physiol. 1979; 63: 375-381Crossref PubMed Google Scholar). Freeze fracture is very useful to address the configuration of biological membranes. However, the definite identification of 10 nm particles remains to be determined because of the limited resolution. In higher plants, two conflicting results on the oligomerization status of PS II are reported using image analyses of two-dimensional crystals (45.Holzenburg A. Bewley M.C. Wilson F.H. Nicholson W.V. Ford R.C. Nature. 1993; 363: 470-472Crossref Scopus (86) Google Scholar, 46.Lyon M.K. Marr K.M. Furcinitti P.S. J. Struct. Biol. 1993; 110: 133-140Crossref PubMed Scopus (52) Google Scholar), which were also obtained by use of detergents. There are also many reports that show the PS II dimer on BN-PAGE (e.g. Refs. 11.Aro E.M. Suorsa M. Rokka A. Allahverdiyeva Y. Paakkarinen V. Saleem A. Battchikova N. Rintamaki E. J. Exp. Bot. 2005; 56: 347-356Crossref PubMed Scopus (380) Google Scholar, 59.Herranen M. Battchikova N. Zhang P. Graf A. Sirpio S. Paakkarinen V. Aro E.M. Plant Physiol. 2004; 134: 470-481Crossref PubMed Scopus (148) Google Scholar, 60.Komenda J. Reisinger V. Muller B.C. Dobakova M. Granvogl B. Eichacker L.A. J. Biol. Chem. 2004; 279: 48620-48629Abstract Full Text Full Text PDF PubMed Scopus (132) Google Scholar). In some reports, higher or comparable amounts of monomer relative to dimer were found, although the authors concluded that the monomer is a transient status of PS II biogenesis (11.Aro E.M. Suorsa M. Rokka A. Allahverdiyeva Y. Paakkarinen V. Saleem A. Battchikova N. Rintamaki E. J. Exp. Bot. 2005; 56: 347-356Crossref PubMed Scopus (380) Google Scholar, 60.Komenda J. Reisinger V. Muller B.C. Dobakova M. Granvogl B. Eichacker L.A. J. Biol. Chem. 2004; 279: 48620-48629Abstract Full Text Full Text PDF PubMed Scopus (132) Google Scholar). The use of detergents to solubilize membrane protein complexes could affect the appearance of the dimer on BN-PAGE, because the concentration of DDM seemed to affect the amount of lipids associated with PS II complexes (Fig. 5). Solubilization at a lower concentration of DDM resulted in diffuse bands, both of PS II and PS I, and some parts of them did not migrate into the resolving gel, although there was no appearance of dimeric PS II (Fig. 5 and supplemental Fig. S3). A more heterogeneous amount of lipids associated with individual PS II complexes solubilized with lower concentrations of DDM could result in the defused band shape. Therefore, aiming to obtain distinct bands of membrane protein complexes, higher concentrations of detergents are used in many reports for BN-PAGE analyses.To exclude the effects of detergents described above when assessing the form of PS II complexes in vivo, we measured photochemical reactions in the absence of detergents using thylakoids and cells of not only of primitive red alga, but also from cyanobacteria and a diatom. Differing from purified PS II complexes, PS I complexes are present and phycobilisomes are attached to PS II complexes in cells and thylakoids in red algae and cyanobacteria. In general, the fluorescence yield of PS I is quite low at room temperature (61.Murata N. Satoh K. Govindjee Amesz J. Fork D.C. Light Emission by Plants and Bacteria. Academic Press, Orlando, FL1986: 137-159Crossref Google Scholar), and the variable fluorescence can be attributed to PS II only. Presence of phycobilisomes can increase the antenna size of PS II, which increases the rate of QA reduction, but cannot change the fluorescence induction pattern unless the PS II core complexes are connected through the large linker protein of the phycobilisome. The presence of phycobilisomes also contributes to the increase of Fo level. In our measurements, energy transfer between PS II units, which can be determined by the sigmoidal fluorescence increase as observed in dimeric PS II, was barely detected in vivo (Fig. 6). This result strongly indicates that most of the population of PS II functions in a monomeric form in vivo. Differing from the materials above, energy transfer between the PS II units was observed in spinach thylakoids and PS II preparations (Fig. 6). This is consistent with the previous reports (44.Joliot A. Joliot P. C. R. Acad. Sci. Paris. 1964; 258: 4622-4625Google Scholar, 47.Melis A. Homann P.H. Photochem. Photobiol. 1976; 23: 343-350Crossref PubMed Scopus (223) Google Scholar, 62.Melis A. Duysens L.N.M. Photochem. Photobiol. 1979; 29: 373-382Crossref Scopus (134) Google Scholar). In this sense, the relationship between individual PS II reaction center units in non-green plants is quite different from that in green plants, whose PS II complexes are assembled in the grana region and are connected directly or via surrounding LHCII.The remaining problem opposing our conclusion is that the oxygen-evolving activity in the PS II monomer is generally lower than in the dimer (e.g. Ref. 39.Sakurai I. Mizusawa N. Wada H. Sato N. Plant Physiol. 2007; 145: 1361-1370Crossref PubMed Scopus (109) Google Scholar and Table 2). However, based on our present investigation, this can be explained by higher amounts of lipids in the monomer, which affect the accessibility of artificial electron acceptors to the secondary electron acceptor QB site (23.Satoh K. Oh-hashi M. Kashino Y. Koike H. Plant Cell Physiol. 1995; 36: 597-605Crossref Scopus (5) Google Scholar). The artificial electron acceptor, 2,5-DCBQ, is known to support a high rate of oxygen evolution by accepting electrons via both the first and the second intrinsic electron acceptor, QA and QB (23.Satoh K. Oh-hashi M. Kashino Y. Koike H. Plant Cell Physiol. 1995; 36: 597-605Crossref Scopus (5) Google Scholar), and is commonly used at ∼0.5 mm to assess oxygen-evolving activity. At lower concentrations, it accepts electrons via QB plastoquinone, but at higher concentrations, it replaces QB plastoquinone and accepts electrons directly from QA, resulting in a lower activity (23.Satoh K. Oh-hashi M. Kashino Y. Koike H. Plant Cell Physiol. 1995; 36: 597-605Crossref Scopus (5) Google Scholar). Another artificial electron acceptor, duroquinone, was shown to accept electrons only via the secondary intrinsic electron acceptor QB, not directly from QA (23.Satoh K. Oh-hashi M. Kashino Y. Koike H. Plant Cell Physiol. 1995; 36: 597-605Crossref Scopus (5) Google Scholar). The hydrophobicity of quinones can be evaluated by the values of CLogP, the calculated logarithm of the partition coefficient; the more hydrophobic a compound is, the greater its CLogP. The CLogP values of 2,5-DCBQ and duroquinone are 1.25 and 2.63, respectively (63.Siraki A.G. Chan T.S. O'Brien P.J. Toxicol. Sci. 2004; 81: 148-159Crossref PubMed Scopus (69) Google Scholar). The difference of Vmax can be interpreted as the difference of the abundance of lipids resulting in the different turnover rates of the artificial electron acceptors at the QB sites. Abundant lipids in the PS II monomer might become a physical barrier for the less hydrophobic 2,5-DCBQ to access to the QB site resulting in a lower Vmax. Inversely, the abundant lipids might assist the accessibility to the QB site for more hydrophobic duroquinone that accept electrons through QB. Therefore, the rate of oxygen evolution can be determined by the accessibility and mobility of 2,5-DCBQ and duroquinone to the QB site and in the lipids around the QB site.PS II complexes are embedded in the lipid layer (thylakoid membrane), therefore our results indicate that most of the population of PS II functions and exists in a monomeric form in vivo. The stoichiometry of ∼1 between isolated PS II and the bound light-harvesting phycobilisome suggested in T. elongatus (64.Kura-Hotta M. Satoh K. Katoh S. Arch. Biochem. Biophys. 1986; 249: 1-7Crossref PubMed Scopus (12) Google Scholar) (formerly Synechococcus sp.) agrees with the monomeric existence of PS II in vivo, although other stoichiometry is also reported in cyanobacterial and red algal cells/thylakoids (65.Cunningham F.X. Dennenberg R.J. Mustardy L. Jursinic P.A. Gantt E. Plant Physiol. 1989; 91: 1179-1187Crossref PubMed Google Scholar, 66.Kursar T.A. Alberte R.S. Plant Physiol. 1983; 72: 409-414Crossref PubMed Google Scholar, 67.Ley A.C. Plant Physiol. 1984; 74: 451-454Crossref PubMed Google Scholar, 68.Ohki K. Okabe Y. Murakami A. Fujita Y. Plant Cell Physiol. 1987; 28: 1219-1226Google Scholar). Our conclusion is important, because it will call for a novel hypotheses for the function of some PS II subunit proteins, the association of antenna pigment-protein complexes, and the processes of assembly and repair of PS II, due to the fact that the current hypotheses stand on the premise that the functional PS II in vivo is a dimer. Furthermore, the oligomerization status of many crystallographic models of membrane protein complexes may need to be re-evaluated, because the protein complexes were purified using processes similar to the purification of PS II complexes. Photosystem II (PS II) 3The abbreviations used are:PS I and PS IIphotosystem I and IIBNblue nativeChlchlorophyllClogPthe calculated logarithm of the partition coefficient2,5-DCBQ2,5-dichloro-p-benzoquinoneDCMU3-(3,4-dichlorophenyl)-1,1-dimethyl ureaDDMn-dodecyl-β-d-maltosideLHCIIlight-harvesting Chl-binding proteinsMES2-(N-morpholino)ethanesulfonic acidLCliquid chromatographyMS/MStandem mass spectrometry. 3The abbreviations used are:PS I and PS IIphotosystem I and IIBNblue nativeChlchlorophyllClogPthe calculated logarithm of the partition coefficient2,5-DCBQ2,5-dichloro-p-benzoquinoneDCMU3-(3,4-dichlorophenyl)-1,1-dimethyl ureaDDMn-dodecyl-β-d-maltosideLHCIIlight-harvesting Chl-binding proteinsMES2-(N-morpholino)ethanesulfonic acidLCliquid chromatographyMS/MStandem mass spectrometry. complexes convert solar energy to biological redox energy. Through this reaction process, water molecules are oxidized and molecular oxygen is released as a byproduct (reviewed in Ref. 1.Renger G. Renger T. Photosynth. Res. 2008; 98: 53-80Crossref PubMed Scopus (243) Google Scholar), which is the only source of molecular oxygen upon which all aerobic organisms on earth rely. PS II core complexes are membrane protein complexes that are composed of >20 distinct subunit proteins and many functional cofactors, including chlorophylls (Chls), carotenoids, plastoquinone, and metal ions (2.Ferreira K.N. Iverson T.M. Maghlaoui K. Barber J. Iwata S. Science. 2004; 303: 1831-1838Crossref PubMed Scopus (2823) Google Scholar, 3.Kamiya N. Shen J.R. Proc. Natl. Acad. Sci. U.S.A. 2003; 100: 98-103Crossref PubMed Scopus (991) Google Scholar, 4.Kashino Y. Lauber W.M. Carroll J.A. Wang Q. Whitmarsh J. Satoh K. Pakrasi H.B. Biochemistry. 2002; 41: 8004-8012Crossref PubMed Scopus (278) Google Scholar, 5.Loll B. Kern J. Saenger W. Zouni A. Biesiadka J. Nature. 2005; 438: 1040-1044Crossref PubMed Scopus (1595) Google Scholar). Similar to many other membrane protein complexes (6.Huang D. Everly R.M. Cheng R.H. Heymann J.B. Schägger H. Sled V. Ohnishi T. Baker T.S. Cramer W.A. Biochemistry. 1994; 33: 4401-4409Crossref PubMed Scopus (104) Google Scholar, 7.Kurisu G. Zhang H. Smith J.L. Cramer W.A. Science. 2003; 302: 1009-1014Crossref PubMed Scopus (582) Google Scholar, 8.Schägger H. Cramer W.A. von Jagow G. Anal. Biochem. 1994; 217: 220-230Crossref PubMed Scopus (1033) Google Scholar, 9.Stroebel D. Choquet Y. Popot J.L. Picot D. Nature. 2003; 426: 413-418Crossref PubMed Scopus (498) Google Scholar, 10.Tsukihara T. Aoyama H. Yamashita E. Tomizaki T. Yamaguchi H. Shinzawa-Itoh K. Nakashima R. Yaono R. Yoshikawa S. Science. 1995; 269: 1069-1674Crossref PubMed Scopus (1286) Google Scholar), two PS II core complexes are believed to associate together to form a homo-dimer with a molecular mass of ∼650 kDa, as shown by crystallographic models (2.Ferreira K.N. Iverson T.M. Maghlaoui K. Barber J. Iwata S. Science. 2004; 303: 1831-1838Crossref PubMed Scopus (2823) Google Scholar, 3.Kamiya N. Shen J.R. Proc. Natl. Acad. Sci. U.S.A. 2003; 100: 98-103Crossref PubMed Scopus (991) Google Scholar, 5.Loll B. Kern J. Saenger W. Zouni A. Biesiadka J. Nature. 2005; 438: 1040-1044Crossref PubMed Scopus (1595) Google Scholar). photosystem I and II blue native chlorophyll the calculated logarithm of the partition coefficient 2,5-dichloro-p-benzoquinone 3-(3,4-dichlorophenyl)-1,1-dimethyl urea n-dodecyl-β-d-maltoside light-harvesting Chl-binding proteins 2-(N-morpholino)ethanesulfonic acid liquid chromatography tandem mass spectrometry. photosystem I and II blue native chlorophyll the calculated logarithm of the partition coefficient 2,5-dichloro-p-benzoquinone 3-(3,4-dichlorophenyl)-1,1-dimethyl urea n-dodecyl-β-d-maltoside light-harvesting Chl-binding proteins 2-(N-morpholino)ethanesulfonic acid liquid chromatography tandem mass spectrometry. The PS II complex turns over dynamically, although it is quite an integrated complex; our current understanding is that the PS II complex that is damaged by high light is disintegrated into a monomeric form and is further dissociated to replace a degraded D1 protein with a de novo synthesized D1 (reviewed in Refs. 11.Aro E.M. Suorsa M. Rokka A. Allahverdiyeva Y. Paakkarinen V. Saleem A. Battchikova N. Rintamaki E. J. Exp. Bot. 2005; 56: 347-356Crossref PubMed Scopus (380) Google Scholar and 12.Nixon P.J. Barker M. Boehm M. de Vries R. Komenda J. J. Exp. Bot. 2005; 56: 357-363Crossref PubMed Scopus (157) Google Scholar). After the replacement, the PS II complex is integrated into a functional form as a dimer. It is supposed that PS II subunit proteins such as PsbI (13.Dobáková M. Tichy M. Komenda J. Plant Physiol. 2007; 145: 1681-1691Crossref PubMed Scopus (65) Google Scholar) or PsbTc (14.Iwai M. Katoh H. Katayama M. Ikeuchi M. Plant Cell Physiol. 2004; 45: 1809-1816Crossref PubMed Scopus (33) Google Scholar) participate in the formation of the PS II dimer. Crystallographic models of PS II have enabled the determination of the accurate molecular architecture of PS II complexes, all of which are in a dimeric form. The most recent crystallographic model of the PS II dimer at 3.0-Å resolution revealed the presence of six detergent molecules located at the interface of the two monomers (5.Loll B. Kern J. Saenger W. Zouni A. Biesiadka J. Nature. 2005; 438: 1040-1044Crossref PubMed Scopus (1595) Google Scholar). Small structural fluctuations during the purification process might allow the invasion of those detergent molecules. However, it is also probable that the PS II complexes exist in the form of a monomer in vivo and the two distinct monomers become a dimer during the purification step incorporating detergents between their interfaces. This idea led us to investigate the actual form of PS II in vivo. Contrary to the above well known dimeric model of a functional PS II core complex, here we show that the PS II core complex functions and exists in a monomeric form in vivo. DISCUSSIONIn this work, we showed that PS II functions mostly in a monomeric form in vivo, contrary to the widely accepted concept that the functional form of PS II complexes is the dimer. This conventional concept is consistent with and stands on the crystallographic models of PS II reported so far (2.Ferreira K.N. Iverson T.M. Maghlaoui K. Barber J. Iwata S. Science. 2004; 303: 1831-1838Crossref PubMed Scopus (2823) Google Scholar, 3.Kamiya N. Shen J.R. Proc. Natl. Acad. Sci. U.S.A. 2003; 100: 98-103Crossref PubMed Scopus (991) Google Scholar, 5.Loll B. Kern J. Saenger W. Zouni A. Biesiadka J. Nature. 2005; 438: 1040-1044Crossref PubMed Scopus (1595) Google Scholar, 40.Kuhl H. Kruip J. Seidler A. Krieger-Liszkay A. Bunker M. Bald D. Scheidig A.J. Rogner M. J. Biol. Chem. 2000; 275: 20652-20659Abstract Full Text Full Text PDF PubMed Scopus (88) Google Scholar, 41.Zouni A. Witt H.T. Kern J. Fromme P. Krauss N. Saenger W. Orth P. Nature. 2001; 409: 739-743Crossref PubMed Scopus (1754) Google Scholar). However, it is natural that the crystallographic models of PS II are dimers, because all of the PS II crystals were made using purified PS II dimers. One prerequisite for obtaining crystals of membrane protein complexes is to remove lipids (26.Kashino Y. J. Chromatogr. B. 2003; 797: 191-216Crossref PubMed Scopus (68) Google Scholar). As we have shown in this work (), the extensive removal of lipids, on purpose or unintentionally, during the purification steps for the crystallization work could result in the high yield of PS II dimer. The side surface opposing the interface between the two monomers in dimeric PS II might be specific to the formation of dimer, because the loss of PsbTc that locates at such an interface results in a decrease in the ratio of dimeric PS II (14.Iwai M. Katoh H. Katayama M. Ikeuchi M. Plant Cell Physiol. 2004; 45: 1809-1816Crossref PubMed Scopus (33) Google Scholar).To our knowledge, no direct evidence on the in vivo form of PS II complexes has been reported. The model of the PS II dimer is primarily based on freeze-fracture electron microscopic studies and crystallographic models. The freeze-fracture studies revealed the presence of pairs of 10 nm particles associated with phycobilisomes, which were suggested to be the dimeric forms of PS II (55.Giddings T.H. Wasmann C. Staehelin L.A. Plant Physiol. 1983; 71: 409-419Crossref PubMed Google Scholar, 56.Giddings Jr., T.H. Staehelin L.A. Biochim. Biophys. Acta. 1979; 546: 373-382Crossref PubMed Scopus (30) Google Scholar, 57.Mörschel E. Schatz G.H. Planta. 1987; 172: 145-154Crossref PubMed Scopus (62) Google Scholar). PS II monomers could associate together to form a dimer in some physiological statuses. Actually, Wollman reported that 10 nm particles observed on the exoplasmic fracture face are attributable to PS II and are apparently able to aggregate once the phycobilisomes are detached in Cyanidium caldarium (58.Wollman F.A. Plant Physiol. 1979; 63: 375-381Crossref PubMed Google Scholar). Freeze fracture is very useful to address the configuration of biological membranes. However, the definite identification of 10 nm particles remains to be determined because of the limited resolution. In higher plants, two conflicting results on the oligomerization status of PS II are reported using image analyses of two-dimensional crystals (45.Holzenburg A. Bewley M.C. Wilson F.H. Nicholson W.V. Ford R.C. Nature. 1993; 363: 470-472Crossref Scopus (86) Google Scholar, 46.Lyon M.K. Marr K.M. Furcinitti P.S. J. Struct. Biol. 1993; 110: 133-140Crossref PubMed Scopus (52) Google Scholar), which were also obtained by use of detergents. There are also many reports that show the PS II dimer on BN-PAGE (e.g. Refs. 11.Aro E.M. Suorsa M. Rokka A. Allahverdiyeva Y. Paakkarinen V. Saleem A. Battchikova N. Rintamaki E. J. Exp. Bot. 2005; 56: 347-356Crossref PubMed Scopus (380) Google Scholar, 59.Herranen M. Battchikova N. Zhang P. Graf A. Sirpio S. Paakkarinen V. Aro E.M. Plant Physiol. 2004; 134: 470-481Crossref PubMed Scopus (148) Google Scholar, 60.Komenda J. Reisinger V. Muller B.C. Dobakova M. Granvogl B. Eichacker L.A. J. Biol. Chem. 2004; 279: 48620-48629Abstract Full Text Full Text PDF PubMed Scopus (132) Google Scholar). In some reports, higher or comparable amounts of monomer relative to dimer were found, although the authors concluded that the monomer is a transient status of PS II biogenesis (11.Aro E.M. Suorsa M. Rokka A. Allahverdiyeva Y. Paakkarinen V. Saleem A. Battchikova N. Rintamaki E. J. Exp. Bot. 2005; 56: 347-356Crossref PubMed Scopus (380) Google Scholar, 60.Komenda J. Reisinger V. Muller B.C. Dobakova M. Granvogl B. Eichacker L.A. J. Biol. Chem. 2004; 279: 48620-48629Abstract Full Text Full Text PDF PubMed Scopus (132) Google Scholar). The use of detergents to solubilize membrane protein complexes could affect the appearance of the dimer on BN-PAGE, because the concentration of DDM seemed to affect the amount of lipids associated with PS II complexes (Fig. 5). Solubilization at a lower concentration of DDM resulted in diffuse bands, both of PS II and PS I, and some parts of them did not migrate into the resolving gel, although there was no appearance of dimeric PS II (Fig. 5 and supplemental Fig. S3). A more heterogeneous amount of lipids associated with individual PS II complexes solubilized with lower concentrations of DDM could result in the defused band shape. Therefore, aiming to obtain distinct bands of membrane protein complexes, higher concentrations of detergents are used in many reports for BN-PAGE analyses.To exclude the effects of detergents described above when assessing the form of PS II complexes in vivo, we measured photochemical reactions in the absence of detergents using thylakoids and cells of not only of primitive red alga, but also from cyanobacteria and a diatom. Differing from purified PS II complexes, PS I complexes are present and phycobilisomes are attached to PS II complexes in cells and thylakoids in red algae and cyanobacteria. In general, the fluorescence yield of PS I is quite low at room temperature (61.Murata N. Satoh K. Govindjee Amesz J. Fork D.C. Light Emission by Plants and Bacteria. Academic Press, Orlando, FL1986: 137-159Crossref Google Scholar), and the variable fluorescence can be attributed to PS II only. Presence of phycobilisomes can increase the antenna size of PS II, which increases the rate of QA reduction, but cannot change the fluorescence induction pattern unless the PS II core complexes are connected through the large linker protein of the phycobilisome. The presence of phycobilisomes also contributes to the increase of Fo level. In our measurements, energy transfer between PS II units, which can be determined by the sigmoidal fluorescence increase as observed in dimeric PS II, was barely detected in vivo (Fig. 6). This result strongly indicates that most of the population of PS II functions in a monomeric form in vivo. Differing from the materials above, energy transfer between the PS II units was observed in spinach thylakoids and PS II preparations (Fig. 6). This is consistent with the previous reports (44.Joliot A. Joliot P. C. R. Acad. Sci. Paris. 1964; 258: 4622-4625Google Scholar, 47.Melis A. Homann P.H. Photochem. Photobiol. 1976; 23: 343-350Crossref PubMed Scopus (223) Google Scholar, 62.Melis A. Duysens L.N.M. Photochem. Photobiol. 1979; 29: 373-382Crossref Scopus (134) Google Scholar). In this sense, the relationship between individual PS II reaction center units in non-green plants is quite different from that in green plants, whose PS II complexes are assembled in the grana region and are connected directly or via surrounding LHCII.The remaining problem opposing our conclusion is that the oxygen-evolving activity in the PS II monomer is generally lower than in the dimer (e.g. Ref. 39.Sakurai I. Mizusawa N. Wada H. Sato N. Plant Physiol. 2007; 145: 1361-1370Crossref PubMed Scopus (109) Google Scholar and Table 2). However, based on our present investigation, this can be explained by higher amounts of lipids in the monomer, which affect the accessibility of artificial electron acceptors to the secondary electron acceptor QB site (23.Satoh K. Oh-hashi M. Kashino Y. Koike H. Plant Cell Physiol. 1995; 36: 597-605Crossref Scopus (5) Google Scholar). The artificial electron acceptor, 2,5-DCBQ, is known to support a high rate of oxygen evolution by accepting electrons via both the first and the second intrinsic electron acceptor, QA and QB (23.Satoh K. Oh-hashi M. Kashino Y. Koike H. Plant Cell Physiol. 1995; 36: 597-605Crossref Scopus (5) Google Scholar), and is commonly used at ∼0.5 mm to assess oxygen-evolving activity. At lower concentrations, it accepts electrons via QB plastoquinone, but at higher concentrations, it replaces QB plastoquinone and accepts electrons directly from QA, resulting in a lower activity (23.Satoh K. Oh-hashi M. Kashino Y. Koike H. Plant Cell Physiol. 1995; 36: 597-605Crossref Scopus (5) Google Scholar). Another artificial electron acceptor, duroquinone, was shown to accept electrons only via the secondary intrinsic electron acceptor QB, not directly from QA (23.Satoh K. Oh-hashi M. Kashino Y. Koike H. Plant Cell Physiol. 1995; 36: 597-605Crossref Scopus (5) Google Scholar). The hydrophobicity of quinones can be evaluated by the values of CLogP, the calculated logarithm of the partition coefficient; the more hydrophobic a compound is, the greater its CLogP. The CLogP values of 2,5-DCBQ and duroquinone are 1.25 and 2.63, respectively (63.Siraki A.G. Chan T.S. O'Brien P.J. Toxicol. Sci. 2004; 81: 148-159Crossref PubMed Scopus (69) Google Scholar). The difference of Vmax can be interpreted as the difference of the abundance of lipids resulting in the different turnover rates of the artificial electron acceptors at the QB sites. Abundant lipids in the PS II monomer might become a physical barrier for the less hydrophobic 2,5-DCBQ to access to the QB site resulting in a lower Vmax. Inversely, the abundant lipids might assist the accessibility to the QB site for more hydrophobic duroquinone that accept electrons through QB. Therefore, the rate of oxygen evolution can be determined by the accessibility and mobility of 2,5-DCBQ and duroquinone to the QB site and in the lipids around the QB site.PS II complexes are embedded in the lipid layer (thylakoid membrane), therefore our results indicate that most of the population of PS II functions and exists in a monomeric form in vivo. The stoichiometry of ∼1 between isolated PS II and the bound light-harvesting phycobilisome suggested in T. elongatus (64.Kura-Hotta M. Satoh K. Katoh S. Arch. Biochem. Biophys. 1986; 249: 1-7Crossref PubMed Scopus (12) Google Scholar) (formerly Synechococcus sp.) agrees with the monomeric existence of PS II in vivo, although other stoichiometry is also reported in cyanobacterial and red algal cells/thylakoids (65.Cunningham F.X. Dennenberg R.J. Mustardy L. Jursinic P.A. Gantt E. Plant Physiol. 1989; 91: 1179-1187Crossref PubMed Google Scholar, 66.Kursar T.A. Alberte R.S. Plant Physiol. 1983; 72: 409-414Crossref PubMed Google Scholar, 67.Ley A.C. Plant Physiol. 1984; 74: 451-454Crossref PubMed Google Scholar, 68.Ohki K. Okabe Y. Murakami A. Fujita Y. Plant Cell Physiol. 1987; 28: 1219-1226Google Scholar). Our conclusion is important, because it will call for a novel hypotheses for the function of some PS II subunit proteins, the association of antenna pigment-protein complexes, and the processes of assembly and repair of PS II, due to the fact that the current hypotheses stand on the premise that the functional PS II in vivo is a dimer. Furthermore, the oligomerization status of many crystallographic models of membrane protein complexes may need to be re-evaluated, because the protein complexes were purified using processes similar to the purification of PS II complexes. In this work, we showed that PS II functions mostly in a monomeric form in vivo, contrary to the widely accepted concept that the functional form of PS II complexes is the dimer. This conventional concept is consistent with and stands on the crystallographic models of PS II reported so far (2.Ferreira K.N. Iverson T.M. Maghlaoui K. Barber J. Iwata S. Science. 2004; 303: 1831-1838Crossref PubMed Scopus (2823) Google Scholar, 3.Kamiya N. Shen J.R. Proc. Natl. Acad. Sci. U.S.A. 2003; 100: 98-103Crossref PubMed Scopus (991) Google Scholar, 5.Loll B. Kern J. Saenger W. Zouni A. Biesiadka J. Nature. 2005; 438: 1040-1044Crossref PubMed Scopus (1595) Google Scholar, 40.Kuhl H. Kruip J. Seidler A. Krieger-Liszkay A. Bunker M. Bald D. Scheidig A.J. Rogner M. J. Biol. Chem. 2000; 275: 20652-20659Abstract Full Text Full Text PDF PubMed Scopus (88) Google Scholar, 41.Zouni A. Witt H.T. Kern J. Fromme P. Krauss N. Saenger W. Orth P. Nature. 2001; 409: 739-743Crossref PubMed Scopus (1754) Google Scholar). However, it is natural that the crystallographic models of PS II are dimers, because all of the PS II crystals were made using purified PS II dimers. One prerequisite for obtaining crystals of membrane protein complexes is to remove lipids (26.Kashino Y. J. Chromatogr. B. 2003; 797: 191-216Crossref PubMed Scopus (68) Google Scholar). As we have shown in this work (), the extensive removal of lipids, on purpose or unintentionally, during the purification steps for the crystallization work could result in the high yield of PS II dimer. The side surface opposing the interface between the two monomers in dimeric PS II might be specific to the formation of dimer, because the loss of PsbTc that locates at such an interface results in a decrease in the ratio of dimeric PS II (14.Iwai M. Katoh H. Katayama M. Ikeuchi M. Plant Cell Physiol. 2004; 45: 1809-1816Crossref PubMed Scopus (33) Google Scholar). To our knowledge, no direct evidence on the in vivo form of PS II complexes has been reported. The model of the PS II dimer is primarily based on freeze-fracture electron microscopic studies and crystallographic models. The freeze-fracture studies revealed the presence of pairs of 10 nm particles associated with phycobilisomes, which were suggested to be the dimeric forms of PS II (55.Giddings T.H. Wasmann C. Staehelin L.A. Plant Physiol. 1983; 71: 409-419Crossref PubMed Google Scholar, 56.Giddings Jr., T.H. Staehelin L.A. Biochim. Biophys. Acta. 1979; 546: 373-382Crossref PubMed Scopus (30) Google Scholar, 57.Mörschel E. Schatz G.H. Planta. 1987; 172: 145-154Crossref PubMed Scopus (62) Google Scholar). PS II monomers could associate together to form a dimer in some physiological statuses. Actually, Wollman reported that 10 nm particles observed on the exoplasmic fracture face are attributable to PS II and are apparently able to aggregate once the phycobilisomes are detached in Cyanidium caldarium (58.Wollman F.A. Plant Physiol. 1979; 63: 375-381Crossref PubMed Google Scholar). Freeze fracture is very useful to address the configuration of biological membranes. However, the definite identification of 10 nm particles remains to be determined because of the limited resolution. In higher plants, two conflicting results on the oligomerization status of PS II are reported using image analyses of two-dimensional crystals (45.Holzenburg A. Bewley M.C. Wilson F.H. Nicholson W.V. Ford R.C. Nature. 1993; 363: 470-472Crossref Scopus (86) Google Scholar, 46.Lyon M.K. Marr K.M. Furcinitti P.S. J. Struct. Biol. 1993; 110: 133-140Crossref PubMed Scopus (52) Google Scholar), which were also obtained by use of detergents. There are also many reports that show the PS II dimer on BN-PAGE (e.g. Refs. 11.Aro E.M. Suorsa M. Rokka A. Allahverdiyeva Y. Paakkarinen V. Saleem A. Battchikova N. Rintamaki E. J. Exp. Bot. 2005; 56: 347-356Crossref PubMed Scopus (380) Google Scholar, 59.Herranen M. Battchikova N. Zhang P. Graf A. Sirpio S. Paakkarinen V. Aro E.M. Plant Physiol. 2004; 134: 470-481Crossref PubMed Scopus (148) Google Scholar, 60.Komenda J. Reisinger V. Muller B.C. Dobakova M. Granvogl B. Eichacker L.A. J. Biol. Chem. 2004; 279: 48620-48629Abstract Full Text Full Text PDF PubMed Scopus (132) Google Scholar). In some reports, higher or comparable amounts of monomer relative to dimer were found, although the authors concluded that the monomer is a transient status of PS II biogenesis (11.Aro E.M. Suorsa M. Rokka A. Allahverdiyeva Y. Paakkarinen V. Saleem A. Battchikova N. Rintamaki E. J. Exp. Bot. 2005; 56: 347-356Crossref PubMed Scopus (380) Google Scholar, 60.Komenda J. Reisinger V. Muller B.C. Dobakova M. Granvogl B. Eichacker L.A. J. Biol. Chem. 2004; 279: 48620-48629Abstract Full Text Full Text PDF PubMed Scopus (132) Google Scholar). The use of detergents to solubilize membrane protein complexes could affect the appearance of the dimer on BN-PAGE, because the concentration of DDM seemed to affect the amount of lipids associated with PS II complexes (Fig. 5). Solubilization at a lower concentration of DDM resulted in diffuse bands, both of PS II and PS I, and some parts of them did not migrate into the resolving gel, although there was no appearance of dimeric PS II (Fig. 5 and supplemental Fig. S3). A more heterogeneous amount of lipids associated with individual PS II complexes solubilized with lower concentrations of DDM could result in the defused band shape. Therefore, aiming to obtain distinct bands of membrane protein complexes, higher concentrations of detergents are used in many reports for BN-PAGE analyses. To exclude the effects of detergents described above when assessing the form of PS II complexes in vivo, we measured photochemical reactions in the absence of detergents using thylakoids and cells of not only of primitive red alga, but also from cyanobacteria and a diatom. Differing from purified PS II complexes, PS I complexes are present and phycobilisomes are attached to PS II complexes in cells and thylakoids in red algae and cyanobacteria. In general, the fluorescence yield of PS I is quite low at room temperature (61.Murata N. Satoh K. Govindjee Amesz J. Fork D.C. Light Emission by Plants and Bacteria. Academic Press, Orlando, FL1986: 137-159Crossref Google Scholar), and the variable fluorescence can be attributed to PS II only. Presence of phycobilisomes can increase the antenna size of PS II, which increases the rate of QA reduction, but cannot change the fluorescence induction pattern unless the PS II core complexes are connected through the large linker protein of the phycobilisome. The presence of phycobilisomes also contributes to the increase of Fo level. In our measurements, energy transfer between PS II units, which can be determined by the sigmoidal fluorescence increase as observed in dimeric PS II, was barely detected in vivo (Fig. 6). This result strongly indicates that most of the population of PS II functions in a monomeric form in vivo. Differing from the materials above, energy transfer between the PS II units was observed in spinach thylakoids and PS II preparations (Fig. 6). This is consistent with the previous reports (44.Joliot A. Joliot P. C. R. Acad. Sci. Paris. 1964; 258: 4622-4625Google Scholar, 47.Melis A. Homann P.H. Photochem. Photobiol. 1976; 23: 343-350Crossref PubMed Scopus (223) Google Scholar, 62.Melis A. Duysens L.N.M. Photochem. Photobiol. 1979; 29: 373-382Crossref Scopus (134) Google Scholar). In this sense, the relationship between individual PS II reaction center units in non-green plants is quite different from that in green plants, whose PS II complexes are assembled in the grana region and are connected directly or via surrounding LHCII. The remaining problem opposing our conclusion is that the oxygen-evolving activity in the PS II monomer is generally lower than in the dimer (e.g. Ref. 39.Sakurai I. Mizusawa N. Wada H. Sato N. Plant Physiol. 2007; 145: 1361-1370Crossref PubMed Scopus (109) Google Scholar and Table 2). However, based on our present investigation, this can be explained by higher amounts of lipids in the monomer, which affect the accessibility of artificial electron acceptors to the secondary electron acceptor QB site (23.Satoh K. Oh-hashi M. Kashino Y. Koike H. Plant Cell Physiol. 1995; 36: 597-605Crossref Scopus (5) Google Scholar). The artificial electron acceptor, 2,5-DCBQ, is known to support a high rate of oxygen evolution by accepting electrons via both the first and the second intrinsic electron acceptor, QA and QB (23.Satoh K. Oh-hashi M. Kashino Y. Koike H. Plant Cell Physiol. 1995; 36: 597-605Crossref Scopus (5) Google Scholar), and is commonly used at ∼0.5 mm to assess oxygen-evolving activity. At lower concentrations, it accepts electrons via QB plastoquinone, but at higher concentrations, it replaces QB plastoquinone and accepts electrons directly from QA, resulting in a lower activity (23.Satoh K. Oh-hashi M. Kashino Y. Koike H. Plant Cell Physiol. 1995; 36: 597-605Crossref Scopus (5) Google Scholar). Another artificial electron acceptor, duroquinone, was shown to accept electrons only via the secondary intrinsic electron acceptor QB, not directly from QA (23.Satoh K. Oh-hashi M. Kashino Y. Koike H. Plant Cell Physiol. 1995; 36: 597-605Crossref Scopus (5) Google Scholar). The hydrophobicity of quinones can be evaluated by the values of CLogP, the calculated logarithm of the partition coefficient; the more hydrophobic a compound is, the greater its CLogP. The CLogP values of 2,5-DCBQ and duroquinone are 1.25 and 2.63, respectively (63.Siraki A.G. Chan T.S. O'Brien P.J. Toxicol. Sci. 2004; 81: 148-159Crossref PubMed Scopus (69) Google Scholar). The difference of Vmax can be interpreted as the difference of the abundance of lipids resulting in the different turnover rates of the artificial electron acceptors at the QB sites. Abundant lipids in the PS II monomer might become a physical barrier for the less hydrophobic 2,5-DCBQ to access to the QB site resulting in a lower Vmax. Inversely, the abundant lipids might assist the accessibility to the QB site for more hydrophobic duroquinone that accept electrons through QB. Therefore, the rate of oxygen evolution can be determined by the accessibility and mobility of 2,5-DCBQ and duroquinone to the QB site and in the lipids around the QB site. PS II complexes are embedded in the lipid layer (thylakoid membrane), therefore our results indicate that most of the population of PS II functions and exists in a monomeric form in vivo. The stoichiometry of ∼1 between isolated PS II and the bound light-harvesting phycobilisome suggested in T. elongatus (64.Kura-Hotta M. Satoh K. Katoh S. Arch. Biochem. Biophys. 1986; 249: 1-7Crossref PubMed Scopus (12) Google Scholar) (formerly Synechococcus sp.) agrees with the monomeric existence of PS II in vivo, although other stoichiometry is also reported in cyanobacterial and red algal cells/thylakoids (65.Cunningham F.X. Dennenberg R.J. Mustardy L. Jursinic P.A. Gantt E. Plant Physiol. 1989; 91: 1179-1187Crossref PubMed Google Scholar, 66.Kursar T.A. Alberte R.S. Plant Physiol. 1983; 72: 409-414Crossref PubMed Google Scholar, 67.Ley A.C. Plant Physiol. 1984; 74: 451-454Crossref PubMed Google Scholar, 68.Ohki K. Okabe Y. Murakami A. Fujita Y. Plant Cell Physiol. 1987; 28: 1219-1226Google Scholar). Our conclusion is important, because it will call for a novel hypotheses for the function of some PS II subunit proteins, the association of antenna pigment-protein complexes, and the processes of assembly and repair of PS II, due to the fact that the current hypotheses stand on the premise that the functional PS II in vivo is a dimer. Furthermore, the oligomerization status of many crystallographic models of membrane protein complexes may need to be re-evaluated, because the protein complexes were purified using processes similar to the purification of PS II complexes. We thank Drs. Tsuneyoshi Kuroiwa, Terry M. Bricker, and Jian-Ren Shen for providing Cyanidioschyzon merolae, the HT-3 strain, and Thermosynechococcus vulcanus, respectively, and Yukari Yonekura and Dr. Shimpei Aikawa for technical assistance. Supplementary Material Download .doc (1.72 MB) Help with doc files Download .doc (1.72 MB) Help with doc files
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