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Dissipating energy by carotenoids

The demonstration of excitation energy dissipation via energy transfer in a cyanobacterial chlorophyll-carotenoid membrane complex provides evidence that this mechanism may also operate in the light-harvesting complex antennae of higher plants.

Diana Kirilovsky

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© 2015 Nature America, Inc. All rights reserved.

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ight is essential for photosynthetic organisms, but when the energy arriving to the photochemical centers exceeds the capacity of energy consumption by cellular metabolism, deleterious side reactions producing reactive oxygen species increase. Plants, algae and cyanobacteria are exposed to progressive and abrupt changes in solar light intensities and therefore rely on protective mechanisms to balance energy flux. One such mechanism involves converting the light-harvesting antennae into energy thermal dissipation complexes. In plants and green algae, the lightcollecting antennae are composed of integral membrane proteins of the light-harvesting complex (LHC) superfamily, which bind chlorophyll (Chl) and carotenoid molecules. The location and orientation of Chls and carotenoids in the protein enable the efficient transfer of light energy into the photosynthetic reaction centers under low irradiance. High irradiance induces a switch from the light-harvesting state to an energy-quenching state. Although it is clear that carotenoids are the principal actors in energy dissipation, no consensus has been reached on the underlying molecular mechanisms. Excitonic interaction between Chl and carotenoid molecules1, electron transfer2 and energy transfer from a Chl excited state to a carotenoid3 were suggested as possible quenching mechanisms, but the existence of the latter has recently been questioned4. Staleva et al.5 now report such a quenching process involving Chl-to-carotenoid energy transfer in high light–induced proteins (Hlips) from cyanobacteria, which suggests that a similar process may be possible in plants. The small cyanobacterial single-helix Hlips, which have sequence similarities with the first and third helix of plant LHC, are considered to be the ancestors of the whole LHC superfamily6,7 but are not involved in light harvesting and energy transfer to photochemical centers in cyanobacteria. Although their exact role in cells is mostly unknown, it has been proposed that they have a photoprotective role in processes 242

involving Chl molecules, such as the synthesis of Chl-binding proteins8, Chl recycling9 and PSII assembly10. To clarify the function of Hlips in photoprotection, Staleva et al.5 studied one of these proteins, named HliD, to which a photoprotective role in the synthesis and assembly of the photosystem II (PSII) has been assigned11. They demonstrated that HliD binds Chl-a and β-carotene with a 3:1 ratio. This stoichiometry is only possible if HliD forms dimers (which is consistent with the molecular weight of isolated complexes) containing six Chl-a and two β-carotenes. A model is proposed in which the HliD dimer adopts the same structural conformation as helices 1 and 3 of the LHC (Fig. 1a). The complex displayed almost no fluorescence, indicating that it efficiently quenches Chl fluorescence. To elucidate the underlying molecular mechanism of energy dissipation, the authors performed femtosecond transient absorption spectroscopy to measure the Chl-a lifetime in this complex. Chl fluorescence decreased more than 50% in less than 100 ps, confirming efficient quenching of excited Chl (Chl*; Fig. 1b). They followed changes of the whole a

fluorescence spectra (450–720 nm) across time and performed a global fitting of these data. The results obtained after Chl-a excitation at 620 nm demonstrated that Chl-a quenching occurs via energy transfer from the excited Chl-a state to the S1 state of the β-carotene in HliD (Fig. 1b). A clear signature of the excited carotenoid S1 appeared after Chl-a excitation. In addition, other pathways of energy dissipation including electron transfer from Chl-a to β-carotene and excitonic coupling were excluded by the absence of nearinfrared absorbance of carotenoid cation and the slow rise (2.1 ps) of the S1 signal, respectively. The work of Staleva et al.5 is crucial in the elucidation of the role of Hlips in cyanobacteria. For the first time, it was clearly demonstrated that Hlips bind Chl-a and carotenoid molecules and that they have a capacity to dissipate excitation energy via energy transfer from Chl to carotenoid. This study also suggests that a similar mechanism may be relevant for energy quenching in eukaryotic components of the LHC superfamily. In plants and algae, in addition to the LHC

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Car*- S1 Heat

Chl

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Figure 1 | Structures and mechanisms in energy dissipation by HliD. (a) Structural model of the cyanobacterial HliD dimer, highlighting the binding of two β-carotenes (orange) and six Chls (positions highlighted in green), which facilitate energy transfer from Chl to carotenoids (Car). (b) Energy dissipation occurs via energy transfer from excited Chl molecules to the excited S1 state of a β-carotene, and an analogous mechanism may also operate in other proteins of the LHC family, including the LHC antennae. nature chemical biology | VOL 11 | April 2015 | www.nature.com/naturechemicalbiology

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antenna, the LHC superfamily includes single-, double- and triple-helix proteins that, like Hlips, do not contribute to light harvesting and have only photoprotective functions. Thus, it is intriguing to speculate that these permanently quenched proteins may dissipate excitation energy via Chl to carotenoid energy transfer. This may also be true for the LHC antennae, which can be considered as derivatives of these fixed quenching proteins. Structural changes appearing during evolution could convert the permanently quenched complexes into flexible ones. Under low light conditions, they are very efficient antennae and transfer most absorbed energy into the reaction centers; under high light and low pH, the complex

may switch into an energy-dissipative state in which the pigment interactions may be similar to those existing in their Hlip ancestors, enabling excitation energy transfer from an excited Chl to a carotenoid. In conclusion, although the work of Staleva et al.1 does not exclude the possibility that other molecular mechanisms may contribute to excitation energy quenching in photosynthetic membranes, it suggests that the principal quenching mechanism in LHC antennae could be similar to that occurring in HliD: efficient dissipation of the Chl-a excited state energy via the carotenoid S1 state. ■

Paris-Saclay, Gif-sur-Yvette, France. e-mail: [email protected]

Diana Kirilovsky is in the Institut de Biologie Intégrative de la Cellule (I2BC), Campus

Competing financial interests

References

1. Bode, S. et al. Proc. Natl. Acad. Sci. USA 106, 12311–12316 (2009). 2. Holt, N.E. Science 307, 433–436 (2005). 3. Ruban, A.V. et al. Nature 450, 575–578 (2007). 4. Müller, M.G. et al. ChemPhysChem 11, 1289–1296 (2010). 5. Staleva, H. et al. Nat. Chem. Biol. 11, 287–291 (2015). 6. Dolganov, N.A.M. et al. Proc. Natl. Acad. Sci. USA 92, 632–640 (1995). 7. Engelken, J. et al. BMC Evol. Biol. 10, 233 (2010). 8. Chidgey, J.W. et al. Plant Cell 26, 1267–1279 (2014). 9. Vavilin, D. et al. J. Biol. Chem. 282, 37660–37668 (2007). 10. Yao, D. et al. J. Biol. Chem. 282, 267–276 (2007). 11. Knoppová, J. et al. Plant Cell 26, 1200–1212 (2014).

The author declares no competing financial interests.

CARBOHYDRATES

Translation from sticky to sweet

Bacterial translation elongation factor P (EF-P) is essential to overcome ribosome stalling at polyproline stretches during protein synthesis. A new mechanism of EF-P activation, identified in a subset of Bacteria, involves addition of the sugar l-rhamnose to a critical arginine residue.

Michela G Tonetti

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olyproline stretches pose problems during protein synthesis: the pyrrolidine ring of proline may impose structural constrains during the formation of the peptide bond, resulting in ribosome stalling. Inhibition of translation at polyproline stretches can be alleviated by the presence of EF-P in Bacteria or its ortholog eIF5A in Eukarya and Archaea1–3. The mechanisms by which EF-P and eIF5A can overcome the effects of proline-rich sequences are not completely understood; however, several pieces of evidence have demonstrated that both factors need to be post-translationally modified to be functionally active. In particular, a conserved lysine residue in eIF5A is converted to the uncommon amino acid hypusine4 (Fig. 1a). In Escherichia coli EF-P, Lys34 is modified using the lysyltransferase EmpA (also known as YjeA), which adds a β-lysine produced by the aminomutase EmpB (YjeK); Lys34 can also be hydroxylated by EmpC (YfcM), but this modification is not strictly required to stimulate EF-P activity (Fig. 1b). The direct inactivation of EF-P or the loss of these modifications caused by inactivation of EmpA (YjeA) or EmpB (YjeK) causes pleiotropic effects, resulting in impaired growth, cell surface

alterations and reduced virulence1,2. These findings indicate that EF-P and its covalent modifications are essential and suggest that this system, owing to its differences with its eukaryotic counterpart, can be an attractive target for the development of new antimicrobial drugs. However, only 26% of Bacteria encode orthologs for EmpA and EmpB, indicating that alternative mechanisms of EF-P activation may be present. Using a phylogenetic approach, Lassak et al.5 have now identified a subset of bacteria characterized by both the lack of the EmpAB cluster and the substitution of the conserved Lys34 with Arg32 in EF-P. The arginine-containing EF-P is associated with a gene encoding a domain of unknown function (DUF2331) in the same cluster. Using defective mutants, Lassak et al.5 demonstrate that this protein, designated EarP, is functionally linked to EF-P and is essential to alleviate ribosome stalling. In vivo and in vitro experiments have clearly shown that EarP is a glycosyltransferase that uses dTDP-l-rhamnose as substrate, transferring the monosaccharide to Arg32. Thus, glycosylation is a third way to promote synthesis of polyproline-containing proteins (Fig. 1c). Protein glycosylation represents the most common post-translational

nature chemical biology | VOL 11 | april 2015 | www.nature.com/naturechemicalbiology

modification. The structure and function of several N- and O-linked glycans are well known. However, in the past years, several noncanonical types of glycosylation, involving new types of linkages and the use of different monosaccharides, have been discovered. These modifications can have profound effects on protein function, contributing to regulation of activity and structure stabilization, as observed, for instance, in O-linked N-acetylglucosamine6. Glycosylation of arginine residues is an extremely rare type of N-linked modification, and very few examples are reported in the literature. It was first identified in corn amylogenin as arginineβ-glucose7. Two independent reports have recently recognized the bacterial virulence factor NleB as a glycosyltransferase, which transfers GlcNAc to conserved arginine residues of death domain–containing proteins, causing inhibition of death receptor signaling8,9. Modification of Arg32 of EF-P with l-rhamnose by the glycosyltransferase EarP represents a further new type of glycosylation. l-rhamnose is a common sugar in Bacteria, where it is used for the production of surface polysaccharides, lipopolysaccharides and rhamnolipids, all of which are involved in virulence. 243

Photosynthesis: Dissipating energy by carotenoids.

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