Spinach-thylakoid phosphorylation: Studies on the kinetics of changes in photosystem antenna size, spill-over and phosphorylation of light-harvesting chlorophyll ab protein (original) (raw)
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Biochimica et Biophysica Acta (BBA) - Bioenergetics, 1995
An open question regarding the changes of energy distribution between Photosystems (PS) I and II following protein phosphorylation and thylakoid destacking is whether or not excitation energy redirected from PS II antenna is effectively trapped by PS I reaction centers. In this report, we measured the effects in spinach thylakoids of Mg2+-depletion and of phosphorylation at 5 and I mM MgC12 on: (1) the effective absorption cross-sections (o-) of both PS II and PSI determined simultaneously from single-turnover flash saturation curves of Chl a fluorescence and of the absorbance change at 820 nm, respectively; (2) the absolute changes in 77 K fluorescence yields emitted by PS II and PS I; and (3) the quenching of room temperature Chl a fluorescence. In all experiments, we observed complementary changes between O'ps i and O-ps n, °ps i consistently increasing at the expense of O-ps ii. ATP-induced decreases of O'r, s u were 6.9% and 11.2% at 5 and 1 mM MgCI 2, respectively, whereas ~rps I increased by 12% and 18.6% under these conditions. In absence of Mg 2÷, O-ps n and °psi changed respectively by-26.2% and +38.9% relative to thylakoids resuspended in presence of 5 mM MgCI 2. These relative increases of ~rps I are larger than the relative decreases of O'ps it by a factor of 1.5-1.7, probably due to the stoichiometry between PS II and PS I complexes typically found in sun-adapted spinach leaves. Also, we observed that the increases of O-es i correspond closely to the increases of 77 K fluorescence yields emitted at 735 nm by PS I. However, no clear relationship could be detected between the changes of °PS 11 and the quenching of both room temperature and 77 K PS II fluorescence. The reasons for such discrepancy are discussed.
FEBS Letters, 1987
Previous studies have indicated that the reversible phosphorylation of a population of antenna complexes that can donate energy to PS II ('mobile LHC II') plays a regulatory role in the state l-state 2 transition in thylakoid membranes. The relationship of phosphorylated LHC II to the multiple PS II-associated chlorophyll a/b-proteins resolvable on green gels is currently unclear. We have used a high resolution gel system to analyze thylakoids phosphorylated in vitro. The only PS II-associated antenna complex to become phosphorylated is CPII*, indicating that this complex represents the mobile LHC II. The other putative PS II antenna complexes, CP29, CP24, and the new complex designated CP27 which comigrates with CPII, are not phosphorylated and are probably components of the bound 'LHC II' antenna. Thylakoid; State l-state 2 transition; LHC II; CP29; Protein phosphorylation; Chlorophyll u/b-light harvesting complex; (Spinach) Published by Elsevier Science Publishers B. V. (Biomedical Division) 00145793/87/$3.50 0 1987 Federation of European Biochemical Societies
Quantification of photosystem I and II in different parts of the thylakoid membrane from spinach
Biochimica Et Biophysica Acta-bioenergetics, 2004
Electron paramagnetic resonance (EPR) was used to quantify Photosystem I (PSI) and PSII in vesicles originating from a series of welldefined but different domains of the thylakoid membrane in spinach prepared by non-detergent techniques. Thylakoids from spinach were fragmented by sonication and separated by aqueous polymer two-phase partitioning into vesicles originating from grana and stroma lamellae. The grana vesicles were further sonicated and separated into two vesicle preparations originating from the grana margins and the appressed domains of grana (the grana core), respectively. PSI and PSII were determined in the same samples from the maximal size of the EPR signal from P700 + and Y D S , respectively. The following PSI/PSII ratios were found: thylakoids, 1.13; grana vesicles, 0.43; grana core, 0.25; grana margins, 1.28; stroma lamellae 3.10. In a sub-fraction of the stroma lamellae, denoted Y-100, PSI was highly enriched and the PSI/PSII ratio was 13. The antenna size of the respective photosystems was calculated from the experimental data and the assumption that a PSII center in the stroma lamellae (PSIIh) has an antenna size of 100 Chl. This gave the following results: PSI in grana margins (PSIa) 300, PSI (PSIh) in stroma lamellae 214, PSII in grana core (PSIIa) 280. The results suggest that PSI in grana margins have two additional light-harvesting complex II (LHCII) trimers per reaction center compared to PSI in stroma lamellae, and that PSII in grana has four LHCII trimers per monomer compared to PSII in stroma lamellae. Calculation of the total chlorophyll associated with PSI and PSII, respectively, suggests that more chlorophyll (about 10%) is associated with PSI than with PSII. D
Biochemistry, 2007
A mild sonication and phase fractionation method has been used to isolate five regions of the thylakoid membrane in order to characterize the functional lateral heterogeneity of photosynthetic reaction centers and light harvesting complexes. Low-temperature fluorescence and absorbance spectra, absorbance cross-section measurements, and picosecond time-resolved fluorescence decay kinetics were used to determine the relative amounts of photosystem II (PSII) and photosystem I (PSI), to determine the relative PSII antenna size, and to characterize the excited-state dynamics of PSI and PSII in each fraction. Marked progressive increases in the proportion of PSI complexes were observed in the following sequence: grana core (BS), whole grana (B3), margins (MA), stroma lamellae (T3), and purified stromal fraction (Y100). PSII antenna size was drastically reduced in the margins of the grana stack and stroma lamellae fractions as compared to the grana. Picosecond time-resolved fluorescence decay kinetics of PSII were characterized by three exponential decay components in the grana fractions, and were found to have only two decay components with slower lifetimes in the stroma. Results are discussed in the framework of existing models of chloroplast thylakoid membrane lateral heterogeneity and the PSII repair cycle. Kinetic modeling of the PSII fluorescence decay kinetics revealed that PSII populations in the stroma and grana margin fractions possess much slower primary charge separation rates and decreased photosynthetic efficiency when compared to PSII populations in the grana stack. 905 688-1855. Phone: 905 688-5550 ext 3826. ‡ Brock University. § Uppsala University. 1 Abbreviations: PS, photosystem; RC, reaction center; Chl, chlorophyll; P680, primary electron donor in photosystem II; Q A , primary quinone electron acceptor in photosystem II; DAS, decay-associated spectrum; F 0, the minimal fluorescence level associated with photochemically active or "open" reaction centers with an oxidized primary quinone electron acceptor, QA; FM, the maximal level of fluorescence associated with photochemically inactive or "closed" reaction centers with reduced primary quinone electron acceptor, Q A .
Biochimica et Biophysica Acta (BBA) - Bioenergetics, 1983
In osmotically shocked pea chloroplasts illuminated with modulated blue-green light 0ight 2), phosphorylation of the light-harvesting chlorophyll a/b-protein complex (LHCP) accompanies the slow decrease in modulated fluorescence that indicates adaptation to light absorbed predominantly by Photosystem II (State 2). On subsequent additional illumination with continuous far-red light (absorbed predominantly by Photosystem I; light 1) both effects are reversed: modulated chlorophyll fluorescence emission increases (indicating adaptation towards State 1) and LHCP is dephosphorylated. Net phosphorylation and dephosphorylation of LHCP induced by light 2 and excess light 1, respectively, occur on the same time scale as the ATP-dependent chlorophyll fluorescence changes indicative of State 2 and State 1 transitions. The phosphatase inhibitor NaF (10 mM), stimulates the effect of blue-green light on fluorescence and prevents the effect of far-red light. These results provide a demonstration that light of different wavelengths can control excitation energy distribution between the two photosystems via the plastoquinol-activated LHCP phosphorylation mechanism suggested previously (
The Journal of Cell Biology, 1984
The functions of the light-harvesting complex of photosystem II (LHC-II) have been studied using thylakoids from intermittent-light-grown (IML) plants, which are deficient in this complex. These chloroplasts have no grana stacks and only limited lamellar appression in situ. In vitro the thylakoids showed limited but significant Mg2+-induced membrane appression and a clear segregation of membrane particles into such regions. This observation, together with the immunological detection of small quantities of LHC-II apoproteins, suggests that the molecular mechanism of appression may be similar to the more extensive thylakoid stacking seen in normal chloroplasts and involve LHC-II polypeptides directly. To study LHC-II function directly, a sonication-freeze-thaw procedure was developed for controlled insertion of purified LHC-II into IML membranes. Incorporation was demonstrated by density gradient centrifugation, antibody agglutination tests, and freeze-fracture electron microscopy. Th...
Plant Physiology, 1992
The photosystem (PS) II antenna system comprises several biochemically and spectroscopically distinct complexes, including light-harvesting complex 11 (LHCII), chlorophyll-protein complex (CP) 29, CP26, and CP24. LHCII, the most abundant of these, is both structurally and functionally diverse. The photosynthetic apparatus is laterally segregated within the thylakoid membrane into PSI-rich and PSII-rich domains, and the distribution of antenna complexes between these domains has implications for antenna function. We report a detailed analysis of the differences in the polypeptide composition of LHCII, CP29, and CP26 complexes associated with grana and stroma thylakoid fractions from spinach (Spinacia oleracea L.), making use of a very high-resolution denaturing gel system, coupled with immunoblots using monospecific antibodies to identify specific antenna components. We first show that the polypeptide composition of the PSII antenna system is more complex than previously thought. We resolved at least five type I LHCII apoproteins and two to three type 11 LHCII apoproteins. We also resolved at least two apoproteins each for CP29 and CP26. In state 1-adapted grana and stroma thylakoid membranes, the spectrum of LHCII apoproteins is surprisingly similar. However, in addition to overall quantitative differences, we saw subtle but reproducible qualitative differences in the spectrum of LHCII apoproteins in grana and stroma membrane domains, including two forms of the major type 11 apoprotein. The implications of these findings for models of PSII antenna function in spinach are discussed. Most of the Chl in the thylakoid membrane of higher plants and green algae is bound by specialized antenna complexes associated with either PSI or PSII (5, 15, 33). The light-harvesting antenna system of PSII includes several distinct CP3 complexes. The major component, LHCII (designated LHCIIb in Thomber's nomenclature; 27), accounts for approximately half of the total Chl as well as of the total protein in the membrane and has a Chl a/b of about 1.1 (21). The minor PSII antenna complexes CP29, CP26, and CP24 (LHCIIa, LHCIIc, and LHCIId in Thomber's nomenclature, 27; these are products of the genes Lhb4, Lhb5, and Lhb6, respectively, 19) each account for a few percent of the total l Supported by National Institutes of Health grant No. GM22912 to L.A.S.
Biochimica Et Biophysica Acta-bioenergetics, 2008
Phosphorylation-dependent movement of the light-harvesting complex II (LHCII) between photosystem II (PSII) and photosystem I (PSI) takes place in order to balance the function of the two photosystems. Traditionally, the phosphorylatable fraction of LHCII has been considered as the functional unit of this dynamic regulation. Here, a mechanical fractionation of the thylakoid membrane of Spinacia oleracea was performed from leaves both in the phosphorylated state (low light, LL) and in the dephosphorylated state (dark, D) in order to compare the phosphorylation-dependent protein movements with the excitation changes occurring in the two photosystems upon LHCII phosphorylation. Despite the fact that several LHCII proteins migrate to stroma lamellae when LHCII is phosphorylated, no increase occurs in the 77 K fluorescence emitted from PSI in this membrane fraction. On the contrary, such an increase in fluorescence occurs in the grana margin fraction, and the functionally important mobile unit is the PSI-LHCI complex. A new model for LHCII phosphorylation driven regulation of relative PSII/PSI excitation thus emphasises an increase in PSI absorption cross-section occurring in grana margins upon LHCII phosphorylation and resulting from the movement of PSI-LHCI complexes from stroma lamellae and subsequent co-operation with the P-LHCII antenna from the grana. The grana margins probably give a flexibility for regulation of linear and cyclic electron flow in plant chloroplasts.
Plant Physiology and Biochemistry, 2011
Pulse-amplitude-modulated (PAM) chlorophyll fluorescence and photosynthetic oxygen evolution were used to investigate the role of the different amount and organization of light-harvesting complexes of photosystem II (LHCII) in four pea species on the susceptibility of the photosynthetic apparatus to highlight treatment. In this work we analyzed the thylakoid membrane lipid composition of the studied pea plants. A relationship between the structural organization of LHCII proteins, the amount of the main lipid classes and the sensitivity of the photosynthetic apparatus to high-light treatment was found. The results reveal that the photosynthetic apparatus, enriched in oligomeric forms of LHCII concomitant with decreased amount of anionic lipids and increased content of the monogalactosyldiacylglycerol (MGDG), is less sensitive to high light. Our data also suggest that the degree of LHCII oligomerization, as well as the lipid composition do not influence the degree of recovery of the PSII photochemistry after excess light exposure.