The stress-induced SCP/HLIP family of small light-harvesting-like proteins (ScpABCDE) protects Photosystem II from photoinhibitory damages in the cyanobacterium <Emphasis Type="Italic">Synechocystis</Emphasis> sp. PCC 6803

Small CAB-like proteins (SCPs) are single-helix light-harvesting-like proteins found in all organisms performing oxygenic photosynthesis. We investigated the effect of growth in moderate salt stress on these stress-induced proteins in the cyanobacterium Synechocystis sp. PCC 6803 depleted of Photosystem I (PSI), which expresses SCPs constitutively, and compared these cells with a PSI-less/ScpABCDE^? mutant. SCPs, by stabilizing chlorophyll-binding proteins and Photosystem II (PSII) assembly, protect PSII from photoinhibitory damages, and in their absence electrons accumulate and will lead to ROS formation. The presence of 0.2 M NaCl in the growth medium increased the respiratory activity and other PSII electron sinks in the PSI-less/ScpABCDE^? strain. We postulate that this salt-induced effect consumes the excess of PSII-generated electrons, reduces the pressure of the electron transport chain, and thereby prevents ¹O₂ production.     

Synechocystis sp PCC 6803 strains lacking photosystem I and phycobilisome function.

To design an in vivo system allowing detailed analysis of photosystem II (PSII) complexes without significant interference from other pigment complexes, part of the psaAB operon coding for the core proteins of photosystem I (PSI) and part of the apcE gene coding for the anchor protein linking the phycobilisome to the thylakoid membrane were deleted from the genome of the cyanobacterium Synechocystis sp strain PCC 6803. Upon transformation and segregation at low light intensity (5 microE m-2 sec-1), a PSI deletion strain was obtained that is light tolerant and grows reasonably well under photoheterotrophic conditions at 5 microE m-2 sec-1 (doubling time approximately 28 hr). Subsequent inactivation of apcE by an erythromycin resistance marker led to reduction of the phycobilin-to-chlorophyll ratio and to a further decrease in light sensitivity. The resulting PSI-less/apcE- strain grew photoheterotrophically at normal light intensity (50 microE m-2 sec-1) with a doubling time of 18 hr. Deletion of apcE in the wild type resulted in slow photoautotrophic growth. The remaining phycobilins in apcE- strains were inactive in transferring light energy to PSII. Cells of both the PSI-less and PSI-less/apcE- strains had an approximately sixfold enrichment of PSII on a chlorophyll basis and were as active in oxygen evolution (on a per PSII basis) as the wild type at saturating light intensity. Both PSI-less strains described here are highly appropriate both for detailed PSII studies and as background strains to analyze site- and region-directed PSII mutants in vivo.     

The small CAB-like proteins of the cyanobacterium Synechocystis sp. PCC 6803: their involvement in chlorophyll biogenesis for Photosystem II

The five small CAB-like proteins (ScpA-E) of the cyanobacterium Synechocystis sp. PCC 6803 belong to the family of stress-induced light-harvesting-like proteins, but are constitutively expressed in a mutant deficient of Photosystem I (PSI). Using absorption, fluorescence and thermoluminescence measurements this PSI-less strain was compared with a mutant, in which all SCPs were additionally deleted. Depletion of SCPs led to structural rearrangements in Photosystem II (PSII): less photosystems were assembled; and in these, the Q(B) site was modified. Despite the lower amount of PSII, the SCP-deficient cells contained the same amount of phycobilisomes (PBS) as the control. Although the excess PBS were functionally disconnected, their fluorescence was quenched under high irradiance by the activated Orange Carotenoid Protein (OCP). Additionally the amount of OCP, but not of the iron-stress induced protein (isiA), was higher in this SCP-depleted mutant compared with the control. As previously described, the lack of SCPs affects the chlorophyll biosynthesis (Vavilin, D., Brune, D. C., Vermaas, W. (2005) Biochim Biophys Acta 1708, 91-101). We demonstrate that chlorophyll synthesis is required for efficient PSII repair and that it is partly impaired in the absence of SCPs. At the same time, the amount of chlorophyll also seems to influence the expression of ScpC and ScpD.     

Role of the PsbI protein in photosystem II assembly and repair in the cyanobacterium Synechocystis sp. PCC 6803

The involvement of the PsbI protein in the assembly and repair of the photosystem II (PSII) complex has been studied in the cyanobacterium Synechocystis sp. PCC 6803. Analysis of PSII complexes in the wild-type strain showed that the PsbI protein was present in dimeric and monomeric core complexes, core complexes lacking CP43, and in reaction center complexes containing D1, D2, and cytochrome b-559. In addition, immunoprecipitation experiments and the use of a histidine-tagged derivative of PsbI have revealed the presence in the thylakoid membrane of assembly complexes containing PsbI and either the precursor or mature forms of D1. Analysis of PSII assembly in the psbI deletion mutant and in strains lacking PsbI together with other PSII subunits showed that PsbI was not required for formation of PSII reaction center complexes or core complexes, although levels of unassembled D1 were reduced in its absence. However, loss of PsbI led to a dramatic destabilization of CP43 binding within monomeric and dimeric PSII core complexes. Despite the close structural relationship between D1 and PsbI in the PSII complex, PsbI turned over much slower than D1, whereas high light-induced turnover of D1 was accelerated in the absence of PsbI. Overall, our results suggest that PsbI is an early assembly partner for D1 and that it plays a functional role in stabilizing the binding of CP43 in the PSII holoenzyme.     

Gross morphological changes in thylakoid membrane structure are associated with photosystem I deletion in Synechocystis sp. PCC 6803

Cells of Synechocystis sp. PCC 6803 lacking photosystem I (PSI-less) and containing only photosystem II (PSII) or lacking both photosystems I and II (PSI/PSII-less) were compared to wild type (WT) cells to investigate the role of the photosystems in the architecture, structure, and number of thylakoid membranes. All cells were grown at 0.5μmol photons m(-2)s(-1). The lumen of the thylakoid membranes of the WT cells grown at this low light intensity were inflated compared to cells grown at higher light intensity. Tubular as well as sheet-like thylakoid membranes were found in the PSI-less strain at all stages of development with organized regular arrays of phycobilisomes on the surface of the thylakoid membranes. Tubular structures were also found in the PSI/PSII-less strain, but these were smaller in diameter to those found in the PSI-less strain with what appeared to be a different internal structure and were less common. There were fewer and smaller thylakoid membrane sheets in the double mutant and the phycobilisomes were found on the surface in more disordered arrays. These differences in thylakoid membrane structure most likely reflect the altered composition of photosynthetic particles and distribution of other integral membrane proteins and their interaction with the lipid bilayer. These results suggest an important role for the presence of PSII in the formation of the highly ordered tubular structures.     

Enhancement of cyclic electron flow around PSI at high light and its contribution to the induction of non-photochemical quenching of chl fluorescence in intact leaves of tobacco plants

Non-photochemical quenching (NPQ) of Chl fluorescence is a mechanism for dissipating excess photon energy and is dependent on the formation of a DeltapH across the thylakoid membranes. The role of cyclic electron flow around photosystem I (PSI) (CEF-PSI) in the formation of this DeltapH was elucidated by studying the relationships between O2-evolution rate [V(O2)], quantum yield of both PSII and PSI [Phi(PSII) and Phi(PSI)], and Chl fluorescence parameters measured simultaneously in intact leaves of tobacco plants in CO2-saturated air. Although increases in light intensity raised V(O2) and the relative electron fluxes through both PSII and PSI [Phi(PSII) x PFD and Phi(PSI) x PFD] only Phi(PSI) x PFD continued to increase after V(O2) and Phi(PSII) x PFD became light saturated. These results revealed the activity of an electron transport reaction in PSI not related to photosynthetic linear electron flow (LEF), namely CEF-PSI. NPQ of Chl fluorescence drastically increased after Phi(PSII) x PFD became light saturated and the values of NPQ correlated positively with the relative activity of CEF-PSI. At low temperatures, the light-saturation point of Phi(PSII) x PFD was lower than that of Phi(PSI) x PFD and NPQ was high. On the other hand, at high temperatures, the light-dependence curves of Phi(PSII) x PFD and Phi(PSI) x PFD corresponded completely and NPQ was not induced. These results indicate that limitation of LEF induced CEF-PSI, which, in turn, helped to dissipate excess photon energy by driving NPQ of Chl fluorescence.     

Net light-induced oxygen evolution in photosystem I deletion mutants of the cyanobacterium Synechocystis sp. PCC 6803

Oxygenic photosynthesis in cyanobacteria, algae, and plants requires photosystem II (PSII) to extract electrons from H(2)O and depends on photosystem I (PSI) to reduce NADP(+). Here we demonstrate that mixotrophically-grown mutants of the cyanobacterium Synechocystis sp. PCC 6803 that lack PSI (ΔPSI) are capable of net light-induced O(2) evolution in vivo. The net light-induced O(2) evolution requires glucose and can be sustained for more than 30min. Utilizing electron transport inhibitors and chlorophyll a fluorescence measurements, we show that in these mutants PSII is the source of the light-induced O(2) evolution, and that the plastoquinone pool is reduced by PSII and subsequently oxidized by an unidentified electron acceptor that does not involve the plastoquinol oxidase site of the cytochrome b(6)f complex. Moreover, both O(2) evolution and chlorophyll a fluorescence kinetics of the ΔPSI mutants are highly sensitive to KCN, indicating the involvement of a KCN-sensitive enzyme(s). Experiments using (14)C-labeled bicarbonate show that the ΔPSI mutants assimilate more CO(2) in the light compared to the dark. However, the rate of the light-minus-dark CO(2) assimilation accounts for just over half of the net light-induced O(2) evolution rate, indicating the involvement of unidentified terminal electron acceptors. Based on these results we suggest that O(2) evolution in ΔPSI cells can be sustained by an alternative electron transport pathway that results in CO(2) assimilation and that includes PSII, the platoquinone pool, and a KCN-sensitive enzyme.     

Inhibition of chlorophyll biosynthesis at the protochlorophyllide reduction step results in the parallel depletion of Photosystem I and Photosystem II in the cyanobacterium Synechocystis PCC 6803

In most oxygenic phototrophs, including cyanobacteria, two independent enzymes catalyze the reduction of protochlorophyllide to chlorophyllide, which is the penultimate step in chlorophyll (Chl) biosynthesis. One is light-dependent NADPH:protochlorophyllide oxidoreductase (LPOR) and the second type is dark-operative protochlorophyllide oxidoreductase (DPOR). To clarify the roles of both enzymes, we assessed synthesis and accumulation of Chl-binding proteins in mutants of cyanobacterium Synechocystis PCC 6803 that either completely lack LPOR or possess low levels of the active enzyme due to its ectopic regulatable expression. The LPOR-less mutant grew photoautotrophically in moderate light and contained a maximum of 20 % of the wild-type (WT) Chl level. Both Photosystem II (PSII) and Photosystem I (PSI) were reduced to the same degree. Accumulation of PSII was mostly limited by the synthesis of antennae CP43 and especially CP47 as indicated by the accumulation of reaction center assembly complexes. The phenotype of the LPOR-less mutant was comparable to the strain lacking DPOR that also contained <25 % of the wild-type level of PSII and PSI when cultivated under light-activated heterotrophic growth conditions. However, in the latter case, we detected no reaction center assembly complexes, indicating that synthesis was almost completely inhibited for all Chl-proteins, including the D1 and D2 proteins.     

Histidine residue 252 of the Photosystem II D1 polypeptide is involved in a light-induced cross-linking of the polypeptide with the alpha subunit of cytochrome b-559: study of a site-directed mutant of Synechocystis PCC 6803

Properties of the Photosystem II (PSII) complex were examined in the wild-type (control) strain of the cyanobacterium Synechocystis PCC 6803 and its site-directed mutant D1-His252Leu in which the histidine residue 252 of the D1 polypeptide was replaced by leucine. This mutation caused a severe blockage of electron transfer between the PSII electron acceptors Q(A) and Q(B) and largely inhibited PSII oxygen evolving activity. Strong illumination induced formation of a D1-cytochrome b-559 adduct in isolated, detergent-solubilized thylakoid membranes from the control but not the mutant strain. The light-induced generation of the adduct was suppressed after prior modification of thylakoid proteins either with the histidine modifier platinum-terpyridine-chloride or with primary amino group modifiers. Anaerobic conditions and the presence of radical scavengers also inhibited the appearance of the adduct. The data suggest that the D1-cytochrome adduct is the product of a reaction between the oxidized residue His(252) of the D1 polypeptide and the N-terminal amino group of the cytochrome alpha subunit. As the rate of the D1 degradation in the control and mutant strains is similar, formation of the adduct does not seem to represent a required intermediary step in the D1 degradation pathway.     

Organization and activity of photosystems in the mesophyll and bundle sheath chloroplasts of maize

Photosystem I and Photosystem II activities, as well as polypeptide content of chlorophyll (Chl)-protein complexes were analyzed in mesophyll (M) and bundle sheath (BS) chloroplasts of maize (Zea mays L.) growing under moderate and very low irradiance. This paper discusses the application of two techniques: mechanical and enzymatic, for separation of M and BS chloroplasts. The enzymatic isolation method resulted in depletion of polypeptides of oxygen evolving complex (OEC) and alphaCF1 subunit of coupling factor; D1 and D2 polypeptides of PSII were reduced by 50%, whereas light harvesting complex of photosystem II (LHCII) proteins were still detectable. Loss of PSII polypeptides correlated with the decreasing of Chl fluorescence measured at room temperature. Using mechanical isolation of chloroplasts from BS cells, all tested polypeptides could be detected. We found a total lack of O2 evolution in BS chloroplasts, but dichlorophenolindophenol (DCPIP) was photoreduced. PSI activity of chloroplasts isolated from 14- and 28-day-old plants was similar in BS chloroplasts in moderate light (ML), but in low light (LL) it was reduced by about 20%. PSI and PSII activities in M chloroplasts of plants growing in ML decreased with aging of plants. In older LL-grown plants, activities of both photosystems were higher than those observed in chloroplasts from ML-grown plants. We suggest that in BS chloroplasts of maize, PSII complex is assembled typically for the agranal membranes (containing mainly stroma thylakoids) and is able to perform very limited electron transport activity. This in turn suggests the role of PSII for poising the redox state of PSI.     

Chloroplast-encoded PsbT is required for efficient biogenesis of photosystem II complex in the green alga Chlamydomonas reinhardtii

The small hydrophobic polypeptide PsbT is associated with the photosystem II (PSII) reaction center (D1/D2 heterodimer). Here, we report the effect of the deletion of PsbT on the biogenesis of PSII complex during light-induced greening of y-1 mutants of the green alga Chlamydomonas reinhardtii. The y-1 is unable to synthesize chlorophylls in the dark but do so in the light. The dark-grown y-1 cells accumulated no major PSII proteins but a small amount of PsbT. Upon illumination, PsbT was immediately synthesized while chlorophylls, major PSII proteins, and O(2)-evolving activity increased after a 1-h lag. The y-1 cells without PsbT accumulated chlorophylls and PSI protein at a similar rate, whereas the accumulation of PSII complex was specifically retarded during greening. The absence of PsbT did not affect the synthesis of PSII proteins. These results indicate that PsbT is required for the efficient biogenesis of PSII complex.     

Dual role of triplet localization on the accessory chlorophyll in the photosystem II reaction center: photoprotection and photodamage of the D1 protein

Infrared absorption and electron spin resonance studies have shown that the excited triplet state of chlorophyll formed by radical pair recombination in the PSII reaction center is mainly localized on the accessory chlorophyll, which is most probably located in the D1 protein (Chl(1)). This triplet localization plays two contrasting roles, depending on the redox state of Q(A), in the process of acceptor-side photoinhibition of PSII. In the early stage of photoinhibition, in which singly reduced Q(A) is reversibly stabilized, the triplet state of Chl(1) ((3)Chl(1)*) is rapidly quenched (t(1/2) = 2-20 micro s) by the interaction with Q(A)(-), preventing formation of harmful singlet oxygen. In the next inhibitory stage, in which Q(A) is doubly reduced and then irreversibly released from the Q(A) pocket, the lifetime of (3)Chl(1)* becomes longer by more than two orders of magnitude (t(1/2) = 1-3 ms). As a result, singlet oxygen is produced around Chl(1) in the D1 protein, causing damage preferably to the D1 protein, which induces subsequent proteolytic degradation. Thus, (3)Chl(1)* functions as a switch to change from the protective to the degradative phase of the PSII reaction center by sensing either reversible or irreversible inhibited state at the Q(A) site.     

Characterization of carotenoid and chlorophyll photooxidation in photosystem II

Photosystem II (PSII) contains two accessory chlorophylls (Chl(Z), ligated to D1-His118, and Chl(D), ligated to D2-His117), carotenoid (Car), and heme (cytochrome b(559)) cofactors that function as alternate electron donors under conditions in which the primary electron-donation pathway from the O(2)-evolving complex to P680(+) is inhibited. The photooxidation of the redox-active accessory chlorophylls and Car has been characterized by near-infrared (near-IR) absorbance, shifted-excitation Raman difference spectroscopy (SERDS), and electron paramagnetic resonance (EPR) spectroscopy over a range of cryogenic temperatures from 6 to 120 K in both Synechocystis PSII core complexes and spinach PSII membranes. The following key observations were made: (1) only one Chl(+) near-IR band is observed at 814 nm in Synechocystis PSII core complexes, which is assigned to Chl(Z)(+) based on previous spectroscopic studies of the D1-H118Q and D2-H117Q mutants [Stewart, D. H., Cua, A., Chisholm, D. A., Diner, B. A., Bocian, D. F., and Brudvig, G. W. (1998) Biochemistry 37, 10040-10046]; (2) two Chl(+) near-IR bands are observed at 817 and 850 nm in spinach PSII membranes which are formed with variable relative yields depending on the illumination temperature and are assigned to Chl(Z)(+), and Chl(D)(+), respectively; (3) the Chl and Car cation radicals have significantly different stabilities at reduced temperatures with Car(+) decaying much faster; (4) in Synechocystis PSII core complexes, Car(+) decays by recombination with Q(A)(-) and not by Chl(Z)/Chl(D) oxidation, with multiphasic kinetics that are attributed to an ensemble of protein conformers that are trapped as the protein is frozen; and (5) in spinach PSII membranes, Car(+) decays mainly by recombination with Q(A)(-), but also partly by formation of the 850 nm Chl cation radical. The greater stability of Chl(Z)(+) at low temperatures enabled us to confirm that resonance Raman bands previously assigned to Chl(Z)(+) are correctly assigned. In addition, the formation and decay of these cations provide insight into the alternate electron-donation pathways to P680(+).     

Genetic Manipulation of the Cyanobacterium Synechocystis sp. PCC 6803 (Development of Strains Lacking Photosystem I for the Analysis of Mutations in Photosystem II)

We have taken a genetic approach to eliminating the presence of photosystem I (PSI) in site-directed mutants of photosystem II (PSII) in the cyanobacterium Synechocystis sp. PCC 6803. By selecting under light-activated heterotrophic conditions, we have inactivated the psaA-psaB operon encoding the PSI reaction center proteins in cells containing deletions of the three psbA genes. We have also introduced deletions into both copies of psbD in a strain containing a mutation that inactivates psaA (ADK9). These strains, designated D1-/PSI- and D2-/PSI-, may serve as recipient strains for the incorporation of site-directed mutations in either psbA2 or psbD1. The characterization of these cells, which lack both PSI and PSII, is described.     

Oxidative stress and photoinhibition can be separated in the cyanobacterium Synechocystis sp. PCC 6803

Roles of oxidative stress and photoinhibition in high light acclimation were studied using a regulatory mutant of the cyanobacterium Synechocystis sp. PCC 6803. The mutant strain ΔsigCDE contains the stress responsive SigB as the only functional group 2 σ factor. The ?sigCDE strain grew more slowly than the control strain in methyl-viologen-induced oxidative stress. Furthermore, a fluorescence dye detecting H₂O₂, hydroxyl and peroxyl radicals and peroxynitrite, produced a stronger signal in ?sigCDE than in the control strain, and immunological detection of carbonylated residues showed more protein oxidation in ?sigCDE than in the control strain. These results indicate that ?sigCDE suffers from oxidative stress in standard conditions. The oxidative stress may be explained by the findings that ?sigCDE had a low content of glutathione and low amount of Flv3 protein functioning in the Mehler-like reaction. Although ?sigCDE suffers from oxidative stress, up-regulation of photoprotective carotenoids and Flv4, Sll2018, Flv2 proteins protected PSII against light induced damage by quenching singlet oxygen more efficiently in ?sigCDE than in the control strain in visible and in UV-A/B light. However, in UV-C light singlet oxygen is not produced and PSII damage occurred similarly in the ?sigCDE and control strains. According to our results, resistance against the light-induced damage of PSII alone does not lead to high light tolerance of the cells, but in addition efficient protection against oxidative stress would be required.     

A strain of Synechocystis sp. PCC 6803 without photosynthetic oxygen evolution and respiratory oxygen consumption: implications for the study of cyclic photosynthetic electron transport

Cyclic electron transport around photosystem (PS) I is believed to play a role in generation of ATP required for adaptation to stress in cyanobacteria and plants. However, elucidation of the pathway(s) of cyclic electron flow is difficult because of low rates of this electron flow relative to those of linear photosynthetic and respiratory electron transport. We have constructed a strain of Synechocystis sp. PCC 6803 that lacks both PSII and respiratory oxidases and that, consequently, neither evolves nor consumes oxygen. However, this strain is still capable of cyclic electron flow around PSI. The photoheterotrophic growth rate of this strain increased with light intensity up to an intensity of about 25 mumol photons m-2 s-1, supporting the notion that cyclic electron flow contributes to ATP generation in this strain. Indeed, the ATP-generating ability of PSI is demonstrated by the fact that the PSII-less oxidase-less strain is able to grow at much higher salt concentrations than a strain lacking PSI. A quinone electrode was used to measure the redox state of the plastoquinone pool in vivo in the various strains used in this study. In contrast to what is observed in chloroplasts, the plastoquinone pool was rather reduced in darkness and was oxidized in the light. This is in line with significant electron donation by respiratory pathways (NADPH dehydrogenase and particularly succinate dehydrogenase) in darkness. In the light, the pool becomes oxidized due to the presence of much more PSI than PSII. In the oxidase-less strains, the plastoquinone pool was very much reduced in darkness and was oxidized in the light by PSI. Photosystem II activity did not greatly alter the redox state of the plastoquinone pool. The results suggest that cyclic electron flow around PSI can contribute to generation of ATP, and a strain deficient in linear electron transport pathways provides an excellent model for further investigations of cyclic electron flow.     

Impairment of the photosynthetic apparatus by oxidative stress induced by photosensitization reaction of protoporphyrin IX

Treatment with the herbicide acifluorfen-sodium (AF-Na), an inhibitor of protoporphyrinogen oxidase, caused an accumulation of protoporphyrin IX (Proto IX) , light-induced necrotic spots on the cucumber cotyledon within 12-24 h, and photobleaching after 48-72 h of light exposure. Proto IX-sensitized and singlet oxygen ((1)O(2))-mediated oxidative stress caused by AF-Na treatment impaired photosystem I (PSI), photosystem II (PSII) and whole chain electron transport reactions. As compared to controls, the F(v)/F(m) (variable to maximal chlorophyll a fluorescence) ratio of treated samples was reduced. The PSII electron donor NH(2)OH failed to restore the F(v)/F(m) ratio suggesting that the reduction of F(v)/F(m) reflects the loss of reaction center functions. This explanation is further supported by the practically near-similar loss of PSI and PSII activities. As revealed from the light saturation curve (rate of oxygen evolution as a function of light intensity), the reduction of PSII activity was both due to the reduction in the quantum yield at limiting light intensities and impairment of light-saturated electron transport. In treated cotyledons both the Q (due to recombination of Q(A)(-) with S(2)) and B (due to recombination of Q(B)(-) with S(2)/S(3)) band of thermoluminescence decreased by 50% suggesting a loss of active PSII reaction centers. In both the control and treated samples, the thermoluminescence yield of B band exhibited a periodicity of 4 suggesting normal functioning of the S states in centers that were still active. The low temperature (77 K) fluorescence emission spectra revealed that the F(695) band (that originates in CP-47) increased probably due to reduced energy transfer from the CP47 to the reaction center. These demonstrated an overall damage to the PSI and PSII reaction centers by (1)O(2) produced in response to photosensitization reaction of protoporphyrin IX in AF-Na-treated cucumber seedlings.     

Small Cab-like proteins retard degradation of photosystem II-associated chlorophyll in Synechocystis sp. PCC 6803: kinetic analysis of pigment labeling with 15N and 13C

Isotope (Na(15)NO(3), ((15)NH(4))SO(4) or [(13)C]glucose) labeling was used to analyze chlorophyll synthesis and degradation rates in a set of Synechocystis mutants that lacked single or multiple small Cab-like proteins (SCPs), as well as photosystem I or II. When all five small Cab-like proteins were inactivated in the wild-type background, chlorophyll stability was not affected unless the scpABCDE(-) strain was grown at a moderately high light intensity of 100-300 micromol photons m(-2) s(-1). However, the half-life time of chlorophyll was 5-fold shorter in the photosystem I-less/scpABCDE(-) strain than in the photosystem I-less strain even when grown at low light intensity (~3 micromol photons m(-2) s(-1)) (32 +/- 5 and 161 +/- 25 h, respectively). In other photosystem I-less mutants that lacked one to four of the scp genes the chlorophyll lifetime was in between these two values, with the chlorophyll lifetime generally decreasing with an increasing number of inactivated scps. In contrast, the chlorophyll biosynthesis rate was only marginally affected by inactivation of scps except when all five scp genes were deleted. Small Cab-like protein deficiency did not significantly affect photoinhibition or turnover of photosystem II-associated beta-carotene. It is concluded that SCPs do not alter the stability of functional photosystem II complexes but retard the degradation of photosystem II-associated chlorophyll, consistent with the proposed involvement of SCPs in photosystem II re-assembly or/and repair processes by temporarily binding chlorophyll while photosystem II protein components are being replaced.     

Absence of the psbH gene product destabilizes photosystem II complex and bicarbonate binding on its acceptor side in Synechocystis PCC 6803.

The PsbH protein, a small subunit of the photosystem II complex (PSII), was identified as a 6-kDa protein band in the PSII core and subcore (CP47-D1-D2-cyt b-559) from the wild-type strain of the cyanobacterium Synechocystis PCC 6803. The protein was missing in the D1-D2-cytochrome b-559 complex and also in all PSII complexes isolated from IC7, a mutant lacking the psbH gene. The following properties of PSII in the mutant contrasted with those in wild-type: (a) CP47 was released during nondenaturing electrophoresis of the PSII core isolated from IC7; (b) depletion of CO2 resulted in a reversible decrease of the QA- reoxidation rate in the IC7 cells; (c) light-induced decrease in PSII activity, measured as 2,5-dimethyl-benzoquinone-supported Hill reaction, was strongly dependent on the HCO3- concentration in the IC7 cells; and (d) illumination of the IC7 cells lead to an extensive oxidation, fragmentation and cross-linking of the D1 protein. We did not find any evidence for phosphorylation of the PsbH protein in the wild-type strain. The results showed that in the PSII complex of Synechocystis attachment of CP47 to the D1-D2 heterodimer appears weakened and binding of bicarbonate on the PSII acceptor side is destabilized in the absence of the PsbH protein.