The Orange Carotenoid Protein of Cyanobacteria - Carotenoids: Physical, Chemical, and Biological Functions and Properties - page 13

that provide solvent accessibility to the carotenoid. These surface features could be the site of the
interaction of the OCP with other chromophores or proteins.
Protein-protein interactions and protein conformational changes, which may unmask binding
sites, alter surface shape, and induce changes in local electrostatic potential are likely essential
to OCP's NPQ mechanism (Scott et al. 2006, Rakhimberdieva et al. 2007a). Glutaraldehyde and
high concentrations of glycerol and sucrose completely eliminate NPQ formation in Synechocystis
PCC6803 (Scott et al. 2006, Rakhimberdieva et al. 2007a), suggesting that this process must involve
changes in the association or conformation of the proteins (phycobilisome and/or the OCP). This
is of interest in the context of similar experiments on photosensors; dehydration or the addition
of glycerol abolishes the large-scale and long-range protein motions of a plant LOV domain and
affects the formation of the physiological signaling state (Iwata et al. 2007). These experiments also
highlight the participation of internal and surface water molecules in the conformational l uctua-
tions, which are required for large-scale and/or long-range motions of proteins.
The OCP's photoprotective function may rely on its dynamic structure in several ways. A cluster
of highly conserved residues that converge at the interface of the two domains and line the pocket
in which a sucrose molecule was observed in the A. maxima OCP structure, Figures 1.3a and 1.4a.
The positioning of the sucrose molecule is reminiscent of an allosteric effector, as it is situated in a
loop between the two domains of the protein. Furthermore, the binding of the sucrose molecule also
involves the linker connecting the two domains of the OCP; the l exibility of this region could facili-
tate large changes in the disposition of the two domains with respect to each other. For example, if
in the “activated” protein the interface between the two domains was opened with the linker acting
as a hinge, it would increase the surface exposure of the carotenoid.
The crystals of the OCP contained two molecules in the asymmetric unit; these were rei ned
independently including manual i tting of the carotenoid molecule into each protein chain. In both,
the 3 -hydroxyequinenone adopts an all-trans coni guration in the protein, however, with a slight

bowing across its length (the average deviation from all-trans is 16°). In contrast to its conformation
in solution, where both terminal rings are in the s-cis conformation with respect to the conjugated
backbone, the terminal ring of the hECN containing the keto group is locked into an s-trans con-
formation via the hydrogen bonds to Tyr 203 and Trp 290. The absorption of blue light by the caro-
tenoid is a potential trigger that may regulate a mechanism to modulate the protein conformation.
Indeed, upon illumination with blue-green light, the OCP (which appears orange) is photoconverted
to a red active form (Wilson et al. 2008). Resonance Raman spectroscopy and light-induced FTIR
difference spectra demonstrated that light absorbance by the OCP induces structural changes not
only in the carotenoid but also in the protein (Wilson et al. 2008). Upon illumination of the OCP, the
apparent conjugation length of hECN increased by about one conjugated bond, and hECN reaches
a less distorted, more planar structure. Although the hECN is still all-trans in the red form, the
relatively small conformational changes of the carotenoid are sufi cient to induce protein confor-
mational changes due to the locked conformation of the carotenoid in the dark-state structure. This
“activated,” OCP, through interaction with the core of the phycobilisome, could elicit an alteration
of the phycobilisome structure leading to the quenched state. Alternatively, the carotenoid of the
OCP could directly interact with a phycobilin chromophore (most probably the terminal acceptor)
and dissipate the absorbed energy. High blue-light intensities could induce changes that can lower
the energy of the carotenoid S 1 state rendering possible the energy transfer from the terminal accep-
tor of the phycobilisome.
Those residues that are absolutely conserved (129 of 318) in the primary structure of the OCP are
likely candidates for important functional roles. Many of these surround the pigment, as shown in
Figures 1.3a and 1.4b. Side-chain conformations and hydrogen-bonding patterns that may involve
internal water molecules are known to play critical roles in the mechanisms by which other photo-
sensitive proteins function. Light-mediated signaling in the PYP, BLUF, and LOV domains relies
on a conformational change in the protein mediated by changes in hydrogen bonding (Anderson
et al. 2004, Kort et al. 2004, Jung et al. 2006). By analogy, the alteration of hydrogen-bonding

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