Dramatic Domain Rearrangements of the Cyanobacterial Orange Carotenoid Protein upon Photoactivation

Liu, Haijun; Zhang, Hao; Orf, Gregory S.; Lü, Yue; Jiang, Jing; King, Jeremy D.; Wolf, Nathan R.; Gross, Michael L.; Blankenship, Robert E.

doi:10.1021/acs.biochem.6b00013

Cited by 62 publications

(58 citation statements)

References 46 publications

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“…This indicates that the absorption spectrum characteristic for the orange form is a result of the specific protein-chromophore interactions provided by a distinct protein conformation. OCP photoconversion leads to reduction of the protein order (21), partial unfolding (17,19,20), or formation of the socalled molten globule state (24), which is typical for some other photoactive proteins (25). By definition, a molten globule has an increased protein volume (26), which was demonstrated for OCP R by small-angle x-ray scattering (SAXS) and size-exclusion chromatography (17,22).…”

Section: Introductionmentioning

confidence: 98%

“…Although the structure of the red form of OCP is still unknown, recent efforts provided valuable information about photoinduced rearrangements of OCP (15)(16)(17)(18)(19)(20). It appeared that the red carotenoid protein (RCP), which is a spectral and functional analog of OCP R , could be obtained by partial proteolysis of OCP upon removal of its C-terminal domain (CTD) (15).…”

Section: Introductionmentioning

confidence: 99%

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Fluorescent Labeling Preserving OCP Photoactivity Reveals Its Reorganization during the Photocycle

Maksimov

Sluchanko

Mironov

et al. 2017

Biophysical Journal

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Orange carotenoid protein (OCP), responsible for the photoprotection of the cyanobacterial photosynthetic apparatus under excessive light conditions, undergoes significant rearrangements upon photoconversion and transits from the stable orange to the signaling red state. This is thought to involve a 12-Å translocation of the carotenoid cofactor and separation of the N-and C-terminal protein domains. Despite clear recent progress, the detailed mechanism of the OCP photoconversion and associated photoprotection remains elusive. Here, we labeled the OCP of Synechocystis with tetramethylrhodamine-maleimide (TMR) and obtained a photoactive OCP-TMR complex, the fluorescence of which was highly sensitive to the protein state, showing unprecedented contrast between the orange and red states and reflecting changes in protein conformation and the distances from TMR to the carotenoid throughout the photocycle. The OCP-TMR complex was sensitive to the light intensity, temperature, and viscosity of the solvent. Based on the observed Fö rster resonance energy transfer, we determined that upon photoconversion, the distance between TMR (donor) bound to a cysteine in the C-terminal domain and the carotenoid (acceptor) increased by 18 Å , with simultaneous translocation of the carotenoid into the N-terminal domain. Time-resolved fluorescence anisotropy revealed a significant decrease of the OCP rotation rate in the red state, indicating that the light-triggered conversion of the protein is accompanied by an increase of its hydrodynamic radius. Thus, our results support the idea of significant structural rearrangements of OCP, providing, to our knowledge, new insights into the structural rearrangements of OCP throughout the photocycle and a completely novel approach to the study of its photocycle and non-photochemical quenching. We suggest that this approach can be generally applied to other photoactive proteins.

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Section: Introductionmentioning

confidence: 98%

Section: Introductionmentioning

confidence: 99%

Fluorescent Labeling Preserving OCP Photoactivity Reveals Its Reorganization during the Photocycle

Maksimov

Sluchanko

Mironov

et al. 2017

Biophysical Journal

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“…As the peptides eluted, they were admitted into the mass spectrometer via a PicoView Nanospray Source (PV550, New Objective, Inc., Woburn, MA) at a spray voltage of 1.8 kV. The mass spectrometer was operated in the positive-ion mode with standard data-dependent acquisition settings as described previously (Liu et al 2016) with the following modifications: for each precursor-ion scan, the top 15 ions with minimal intensity of 4 × 10 4 counts were selected for fragmentation with an isolation width of 3.0 m/z and normalized collision energy of 30% of maximum. Data processing and analysis was by Protein Prospector, see references cited in Liu et al (2016).…”

Section: Methodsmentioning

confidence: 99%

The proteolysis adaptor, NblA, binds to the N-terminus of β-phycocyanin: Implications for the mechanism of phycobilisome degradation

et al. 2017

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Phycobilisome (PBS) complexes are massive light-harvesting apparati in cyanobacteria that capture and funnel light energy to the photosystem. PBS complexes are dynamically degraded during nutrient deprivation, which causes severe chlorosis, and resynthesized during nutrient repletion. PBS degradation occurs rapidly after nutrient step-down, and is specifically triggered by non-bleaching protein A (NblA), a small proteolysis adaptor that facilitates interactions between a Clp chaperone and phycobiliproteins. Little is known about the mode of action of NblA during PBS degradation. In this study, we used chemical cross-linking coupled with LC-MS/MS to investigate the interactions between NblA and phycobiliproteins. An isotopically-coded BS3 cross-linker captured a protein interaction between NblA and β-phycocyanin (PC). LC-MS/MS analysis identified the amino acid residues participating in the binding reaction, and demonstrated that K52 in NblA is cross-linked to T2 in β-PC. These results were modeled onto the existing crystal structures of NblA and PC by protein-docking simulations. Our data indicate that the C-terminus of NblA fits in an open groove of β-PC, a region located inside the central hollow cavity of a PC rod. NblA may mediate PBS degradation by disrupting the structural integrity of the PC rod from within the rod. In addition, M1-K44 and M1-K52 cross-links between the N-terminus of NblA and the C-terminus of NblA are consistent with the NblA crystal structure, confirming that the purified NblA is structurally biologically relevant. These findings provide direct evidence that NblA physically interacts with β-PC.

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“…Strong blue-green light induces the breaking of the hydrogen bonds between the carotenoid and the protein and those between the CTD and NTD, leading to the opening of the protein and the translocation of the carotenoid toward the NTD. The photoactivated OCP is in its red configuration (OCP r ), with the two domains completely separated and the carotenoid buried in the NTD (Wilson et al, 2012;Liu et al, 2014Liu et al, , 2016Gupta et al, 2015;Leverenz et al, 2015;Maksimov et al, 2017a). In OCP r , NTD interacts with the phycobilisome (PBS) core, the cyanobacterial antenna, inducing an increase of the thermal dissipation of the excess excitation energy absorbed by PBS (Wilson et al, 2012;Leverenz et al, 2014;Harris et al, 2016).…”

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confidence: 99%

Paralogs of the C-Terminal Domain of the Cyanobacterial Orange Carotenoid Protein Are Carotenoid Donors to Helical Carotenoid Proteins

et al. 2017

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The photoactive Orange Carotenoid Protein (OCP) photoprotects cyanobacteria cells by quenching singlet oxygen and excess excitation energy. Its N-terminal domain is the active part of the protein, and the C-terminal domain regulates the activity. Recently, the characteristics of a family of soluble carotenoid-binding proteins (Helical Carotenoid Proteins [HCPs]), paralogs of the N-terminal domain of OCP, were described. Bioinformatics studies also revealed the existence of genes coding for homologs of CTD. Here, we show that the latter genes encode carotenoid proteins (CTDHs). This family of proteins contains two subgroups with distinct characteristics. One CTDH of each clade was further characterized, and they proved to be very good singlet oxygen quenchers. When synthesized in Escherichia coli or Synechocystis PCC 6803, CTDHs formed dimers that share a carotenoid molecule and are able to transfer their carotenoid to apo-HCPs and apo-OCP. The CTDHs from clade 2 have a cysteine in position 103. A disulfide bond is easily formed between the monomers of the dimer preventing carotenoid transfer. This suggests that the transfer of the carotenoid could be redox regulated in clade 2 CTDH. We also demonstrate here that apoOCPs and apo-CTDHs are able to take the carotenoid directly from membranes, while HCPs are unable to do so. HCPs need the presence of CTDH to become holo-proteins. We propose that, in cyanobacteria, the CTDHs are carotenoid donors to HCPs.Photosynthetic organisms performing oxygenic photosynthesis use solar energy, water, and inorganic carbon to produce all the organic molecules they need.Photosynthesis converts the absorbed energy into chemical energy and the reduction power necessary for the assimilation of CO 2 and the synthesis of organic carbon molecules. However, photosynthetic organisms cannot control the incoming flux of light, and too much light generates secondary reactions creating dangerous species of oxygen leading to cellular damage. Thus, in order to survive, photosynthetic organisms have developed a large variety of photoprotective mechanisms. One of them decreases the energy arriving at the reaction centers by increasing the thermal dissipation of excess excitation energy at the level of the antennae (for review, see Niyogi and Truong, 2013). In cyanobacteria, a soluble carotenoid protein, the Orange Carotenoid Protein (OCP), is essential for this mechanism, known as the OCP-related nonphotochemical quenching mechanism (Wilson et al., 2006; for review, see Kirilovsky and Kerfeld, 2016). In addition, OCP has a second photoprotective activity. It is a very good singlet oxygen ( 1 O 2 ) quencher (Kerfeld et al., 2003;Sedoud et al., 2014).OCP is a photoactive soluble carotenoid protein (Wilson et al., 2008). It is composed of two globular domains: an a-helical N-terminal domain (NTD; residues 18-165) that is unique to cyanobacteria and an a-helix/b-sheet C-terminal domain (CTD; residues

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Dramatic Domain Rearrangements of the Cyanobacterial Orange Carotenoid Protein upon Photoactivation

Cited by 62 publications

References 46 publications

Fluorescent Labeling Preserving OCP Photoactivity Reveals Its Reorganization during the Photocycle

Fluorescent Labeling Preserving OCP Photoactivity Reveals Its Reorganization during the Photocycle

The proteolysis adaptor, NblA, binds to the N-terminus of β-phycocyanin: Implications for the mechanism of phycobilisome degradation

Paralogs of the C-Terminal Domain of the Cyanobacterial Orange Carotenoid Protein Are Carotenoid Donors to Helical Carotenoid Proteins

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