Lifetimes of photosystem I and II proteins in the cyanobacterium Synechocystis sp. PCC 6803 (original) (raw)
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Photosystem II Component Lifetimes in the Cyanobacterium Synechocystis sp. Strain PCC 6803
Journal of Biological Chemistry, 2011
Background: We monitored photosystem II (PSII) protein lifetimes with or without small Cab-like proteins (SCPs) by 15 N labeling and mass spectrometry. Results: Without SCPs, nascent PSII complexes were destabilized, and early chlorophyll precursors were reduced. Conclusion: SCPs play a major role in chlorophyll biosynthesis and nascent PSII stabilization. Significance: Proteins temporarily associated with a protein complex may affect biosynthesis of the complex and cofactors. To gain insight in the lifetimes of photosystem II (PSII) chlorophyll and proteins, a combined stable isotope labeling (15 N)/ mass spectrometry method was used to follow both old and new pigments and proteins. Photosystem I-less Synechocystis cells were grown to exponential or post-exponential phase and then diluted in BG-11 medium with [ 15 N]ammonium and [ 15 N]nitrate. PSII was isolated, and the masses of PSII protein fragments and chlorophyll were determined. Lifetimes of PSII components ranged from 1.5 to 40 h, implying that at least some of the proteins and chlorophyll turned over independently from each other. Also, a significant amount of nascent PSII components accumulated in thylakoids when cells were in post-exponential growth phase. In a mutant lacking small Cab-like proteins (SCPs), most PSII protein lifetimes were unaffected, but the lifetime of chlorophyll and the amount of nascent PSII components that accumulated were decreased. In the absence of SCPs, one of the PSII biosynthesis intermediates, the monomeric PSII complex without CP43, was missing. Therefore, SCPs may stabilize nascent PSII protein complexes. Moreover, upon SCP deletion, the rate of chlorophyll synthesis and the accumulation of early tetrapyrrole precursors were drastically reduced. When [ 14 N]aminolevulinic acid (ALA) was supplemented to 15 N-BG-11 cultures, the mutant lacking SCPs incorporated much more exogenous ALA into chlorophyll than the control demonstrating that ALA biosynthesis was impaired in the absence of SCPs. This illustrates the major effects that nonstoichiometric PSII components such as SCPs have on intermediates and assembly but not on the lifetime of PSII proteins.
Function and association of CyanoP in photosystem II of Synechocystis sp. PCC 6803
Research on Chemical Intermediates, 2014
The CyanoP protein is a cyanobacterial homolog of the PsbP protein, which is an extrinsic subunit of photosystem II (PSII) in green plant species. The molecular function of CyanoP has been investigated in mutant strains of Synechocystis but inconsistent results have been reported by different laboratories. In this study, we generated and characterized a Synechocystis mutant in which entire region of the CyanoP gene was eliminated. After repeated subculture in CaCl 2depleted medium, growth retardation was clearly observed for a CyanoP knockout mutant of Synechocystis sp. PCC 6803 (DP). The PSII-mediated oxygen-evolving activity of the DP cells was more susceptible to depletion of CaCl 2 than that of wildtype cells. The 77 K fluorescence emission spectra indicated that energy coupling between phycobilisome and PSII was perturbed in both wild-type and DP cells under CaCl 2-depleted conditions, and was more evident for the DP mutant. To examine the association of CyanoP with PSII complexes, we tested several detergents for solubilization of thylakoid membranes and showed that CyanoP was partly included in fractions containing large protein complexes in gel-filtration analysis. These results indicate that CyanoP constitutively stabilizes PSII functionality in vivo.
European Journal of Biochemistry, 1995
Photosystem I and I1 core complexes were resolved in a single step from the thylakoid menibrane of Synechot.y.rti.v sp. PCC 6803 by using a mild solubilization procedure in dodecyl /~-t)-maltoside arid Deriphat/PAGE. For each photosystem, two green bands were obtained containing oligomcric and monomeric forms of the core complexes of either photosystem. The oligoniers are likcly to be trimei-s i n the case of photosystem T and dinicrs for photosystem 11, The absorption spectra, polypeptide and pigment composition of green bands corresponding to cither photosystem I or photosyslem TI wci-e identical for monomeric and oligomeric forms. Thc cytochromc b-559 content of photosystem I1 was evalualed to bc one cytochrome h-S59/reaction centre both in the monomeric and diineric forms. Two new I5-kDa and 22-kDa carotenoid-binding proteins were isolated and their polypeptides purified to homogeneity. Keywords: photosystem I ; photosystem 11; pigment-protein complexes: cyanobacteria.
The Journal of biological chemistry, 1989
The cyanobacterium Synechocystis sp. PCC 6803 carries out oxygenic photosynthesis analogous to higher plants. Its photosystem I contains seven different polypeptide subunits. The cartridge mutagenesis technique was used to inactivate the psaD gene which encodes subunit II of photosystem I. A mutant strain lacking subunit II was generated by transforming wild type cells with cloned DNA in which psaD gene was interrupted by a gene conferring kanamycin resistance. The photoautotrophic growth of mutant strain is much slower than that of wild type cells. The membranes prepared from mutant cells lack subunit II of photosystem I. Studies on the purified photosystem I reaction center revealed that the complex lacking subunit II is assembled and is functional in P700 photooxidation but at much reduced rate. Therefore, subunit II of photosystem I is required for efficient function of photosystem I.
The Journal of biological chemistry, 1994
The chlorophyll protein organization has been investigated in thylakoid membranes from mutants of the cyanobacterium Synechocystis sp. PCC 6803, in which the photosystem II (PS II) genes psbB and/or psbC (coding for CP47 and CP43, respectively) were inactivated together with the psaAB operon (coding for the photosystem I (PS I) core complex) and the apcE gene (coding for the phycobilisome anchor protein). Lack of the CP43 protein led to a significant decrease of the D1, D2, and CP47 proteins and a decrease in the 77 K fluorescence emission peak at 685 nm. In the absence of the CP47 protein, no PS II reaction center assembly was detected and the 77 K fluorescence emission peak at 695 nm was lost. The psbB-/psbC-/PS I-less/apcE- mutant had no assembly of the D1, D2, CP47, and CP43 proteins, had lost the 77 K fluorescence emission peaks at 685 and 695 nm, but retained about 15% of the chlorophyll present in the PS I-less/apcE- background strain. A broad 77 K fluorescence emission band ...
Plant Physiology, 2007
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 withi...
European Journal of Biochemistry, 2001
A photosystem II (PSII) core complex lacking the internal antenna CP43 protein was isolated from the photosystem II of Synechocystis PCC6803, which lacks photosystem I (PSI). CP47-RC and reaction centre (RCII) complexes were also obtained in a single procedure by direct solubilization of whole thylakoid membranes. The CP47-RC subcore complex was characterized by SDS/PAGE, immunoblotting, MALDI MS, visible and fluorescence spectroscopy, and absorption detected magnetic resonance. The purity and functionality of RCII was also assayed. These preparations may be useful for mutational analysis of PSII RC and CP47 -RC in studying primary reactions of oxygenic photosynthesis.
Photosynthesis Research, 1996
Synechococcus sp. PCC 7942 (Anacystis nidulans R2) contains two forms of the Photosystem II reaction centre protein D1, which differ in 25 of 360 amino acids. DI:I predominates under low light but is transiently replaced by D1:2 upon shifts to higher light. Mutant cells containing only DI:I have lower photochemical energy capture efficiency and decreased resistance to photoinhibition, compared to cells containing Dl:2. We show that when dark-adapted or under low to moderate light, cells with DI:I have higher non-photochemical quenching of PS II fluorescence (higher qN) than do cells with Dl:2. This is reflected in the 77 K chlorophyll emission spectra, with lower Photosystem II fluorescence at 697-698 nm in cells containing DI:I than in cells with Dl:2. This difference in quenching of Photosystem II fluorescence occurs upon excitation of both chlorophyll at 435 nm and phycobilisomes at 570 nm. Measurement of time-resolved room temperature fluorescence shows that Photosystem II fluorescence related to charge stabilization is quenched more rapidly in cells containing DI:I than in those with D1:2. Cells containing D1:1 appear generally shifted towards State II, with PS II down-regulated, while cells with D1:2 tend towards State I. In these cyanobacteria electron transport away from PS II remains non-saturated even under photoinhibitory levels of light. Therefore, the higher activity of D1:2 Photosystem II centres may allow more rapid photochemical dissipation of excess energy into the electron transport chain. D 1:1 confers capacity for extreme State II which may be of benefit under low and variable light. Abbreviations: D 1-t h e atrazine-binding 32 kDa protein of the PS II reaction centre core; D 1:1-the D 1 protein constitutively expressed during acclimated growth in Synechococcus sp. PCC 7942; D 1:2-an alternate form of the D1 protein induced under excess excitation in Synechococcus sp.
Plant Physiology, 1987
psbA in Synechocystis 6803 was found to belong to a small multigene family with three copies. The psbA gene family was inactivated in vitro by insertion of bacterial drug resistance markers. Inactivation of all three genes resulted in a transformant that is unable to grow photosynthetically but can be cultured photoheterotrophically. This mutant lacks oxygen evolving capacity but retains photosystem I activity. Room temperature measurements of chlorophyll a fluorescence induction demonstrated that the transformant exhibits a high fluorescence yield with little or no variable fluorescence. Immunoblot analyses showed complete loss of the psbA gene product (the DI polypeptide) from thylakoid membranes in the transformant. However, the extrinsic 33 kilodalton polypeptide of the water-splitting complex of photosystem II, is still present. The results indicate that assembly of a partial photosystem II complex may occur even in the absence of the intrinsic Dl polypeptide, a protein implicated as a crucial component of the photosystem II reaction center.