Native Mass Spectrometry Analysis of Oligomerization States of Fluorescence Recovery Protein and Orange Carotenoid Protein: Two Proteins Involved in the Cyanobacterial Photoprotection Cycle

Lü, Yue; Liu, Haijun; Saer, Rafael G.; Zhang, Hao; Meyer, Christine; Li, Veronica L.; Shi, Liuqing; King, Jeremy D.; Gross, Michael L.; Blankenship, Robert E.

doi:10.1021/acs.biochem.6b01094

Cited by 29 publications

(42 citation statements)

References 48 publications

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“…2A). A previous ion mobility(IM)-MS analysis shows one intermediate state in the unfolding process of dFRP 22 , and none in the unfolding of the CTD owing to its compact structure 39 . Here, two intermediate states occur during the unfolding of CTD-dFRP, driven by the higher order structure refolding, suggesting high stability of this complex (Fig.…”

Section: Resultsmentioning

confidence: 92%

“…Immunoprecipitation, size-exclusion chromatography (SEC), native-gel electrophoresis, and molecular modelling all suggest that the CTD is the domain that interacts with FRP 16, 21, 47 . Our previous native MS study shows that FRP primarily exists as a dimer 22 , although two different oligomeric states were identified by crystallography 21 .…”

Section: Resultsmentioning

confidence: 99%

“…As a reference, intact OCP was also mixed with FRP in a 1:2 ratio to investigate the affinity of FRP to OCP o . The IM-MS experiment and data processing were carried out as previously described 22 .…”

Section: Methodsmentioning

confidence: 99%

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A Molecular Mechanism for Nonphotochemical Quenching in Cyanobacteria

et al. 2017

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The cyanobacterial Orange Carotenoid Protein (OCP) protects photosynthetic cyanobacteria from photodamage by dissipating excess excitation energy collected by phycobilisomes (PBS) as heat. Dissociation of the PBS-OCP complex in vivo is facilitated by another protein known as the Fluorescence Recovery Protein (FRP), which primarily exists as a dimeric complex. We used various mass spectrometry(MS)-based techniques to investigate the molecular mechanism of this FRP-mediated process. FRP in the dimeric state (dFRP) retains its high affinity to the C-terminal domain (CTD) of OCP in the red state (OCPr). Site-directed mutagenesis and native MS suggest the head region on FRP is a binding candidate to OCP. After attachment to the CTD, the conformational changes of dFRP enable dFRP to bridge the two domains, facilitating the reversion of OCPr into the orange state (OCPo) accompanied by a structural rearrangement of dFRP. Interestingly, we found a mutual response between FRP and OCP; that is, FRP and OCPr destabilize each other, whereas FRP and OCPo stabilize each other. A detailed mechanism of FRP function is proposed on the basis of the experimental results.

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Section: Resultsmentioning

confidence: 92%

Section: Resultsmentioning

confidence: 99%

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A Molecular Mechanism for Nonphotochemical Quenching in Cyanobacteria

et al. 2017

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“…FRP affinity to OCP r is strong, and its presence during illumination decreases the amplitude of OCP-triggered PBS fluorescence quenching by accelerating the OCP r -to-OCP o conversion (Gwizdala et al, 2011(Gwizdala et al, , 2013Kuzminov et al, 2012;Sluchanko et al, 2017;Thurotte et al, 2017) and maybe also by interacting with OCP o (with low affinity) and preventing its photoactivation . FRP is present mostly as a dimer in solution (Sutter et al, 2013;Lu et al, 2017;Sluchanko et al, 2017). It was proposed that it could monomerize upon binding to CTD .…”

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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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“…These contradictory results raised the doubt that the OCP-dimer is might be an artefact of the high concentration of protein used for crystallization. Moreover, at higher protein concentration the presence of dimeric form of OCP is as compared to monomeric form (Lu et al 2016). What is the real functionally relevant state of OCP is, therefore, still an open question?…”

Section: Trigger: Photo-conversion Of Ocp O To Ocp Rmentioning

confidence: 99%

Site, trigger, quenching mechanism and recovery of non-photochemical quenching in cyanobacteria: recent updates

Sonani¹,

Gardiner²,

Rastogi³

et al. 2018

Photosynth Res

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Cyanobacteria exhibit a novel form of non-photochemical quenching (NPQ) at the level of the phycobilisome. NPQ is a process that protects photosystem II (PSII) from possible highlight-induced photo-damage. Although significant advancement has been made in understanding the NPQ, there are still some missing details. This critical review focuses on how the orange carotenoid protein (OCP) and its partner fluorescence recovery protein (FRP) control the extent of quenching. What is and what is not known about the NPQ is discussed under four subtitles; where does exactly the site of quenching lie? (site), how is the quenching being triggered? (trigger), molecular mechanism of quenching (quenching) and recovery from quenching. Finally, a recent working model of NPQ, consistent with recent findings, is been described.

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Native Mass Spectrometry Analysis of Oligomerization States of Fluorescence Recovery Protein and Orange Carotenoid Protein: Two Proteins Involved in the Cyanobacterial Photoprotection Cycle

Cited by 29 publications

References 48 publications

A Molecular Mechanism for Nonphotochemical Quenching in Cyanobacteria

A Molecular Mechanism for Nonphotochemical Quenching in Cyanobacteria

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

Site, trigger, quenching mechanism and recovery of non-photochemical quenching in cyanobacteria: recent updates

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