Raman Optical Activity Reveals Carotenoid Photoactivation Events in the Orange Carotenoid Protein in Solution

Abstract

The orange carotenoid protein (OCP) plays an important role in photoprotection in cyanobacteria, which is achieved by the photoconversion from the orange dark state (OCP) to the red active state (OCP). Using Raman optical activity (ROA), we studied the conformations of the carotenoid chromophore in the active sites of OCP and OCP. This ROA measurement directly observed the chromophore conformation of native OCP in solution, and the measurement of OCP first demonstrated the ROA spectroscopy for the transient species. For OCP, the spectral features of ROA were mostly reproduced by the quantum chemical calculation based on the crystal structure of the OCP. Within the spatial resolution (∼2 Å), a slight modification of the polyene-chain distortion improved the agreement between the observed and calculated ROA spectra. While the crystal structure of OCP is not available, the ROA spectrum of OCP was reproduced by using the crystal structure of red carotenoid protein (RCP), an OCP proxy. The present results showed that the chromophore conformations in the crystal structures of OCP and RCP hold true for OCP and OCP in solution. Particularly, ROA spectroscopy of the native OCP provides a direct support for the 12 Å translocation of chromophore in the photoactivation, which was proposed by X-ray crystallography using RCP [R. L. Leverenz, M. Sutter, et al. Science 2015, 348, 1463-1466].

Resonance Raman optical activity of the imidazole–Myoglobin complex: Titrating enhancement

“…No secondary structure fingerprint could be identified in the 532‐nm excited rROA spectra of met‐Mb nor imidazolylmet‐Mb, because the resonance with the Q absorption bands of the haem chromophore (see Figure ) led to the selective enhancement of the porphyrin vibrations . On top of that, the concentration of 0.284‐mM Mb (5 mg ml −1 ) used for the measurements was too low to actually detect signals from the globin backbone, further confirming the nature of the rROA signals . The main challenge to understanding the spectral trends currently is to link the conformational modification occurring at the chromophore during ligand binding with the rROA spectral changes.…”

“…In the rRaman spectral region, around 1,000 cm −1 hydrogen‐out‐of‐plane modes can be observed. Fujisawa et al reported in their study on the orange carotenoid protein that the intensity of the rROA band at 980 cm −1 represents a good estimation of the distortion of the chromophore . Despite the structure of the porphyrin differs substantially from that one of the orange carotenoid protein and a direct comparison between the two molecules cannot be done, the lack of good S/N details in the ROA spectrum of imidazolyl‐metMb in correspondence of the Raman bands at 1,002 and 976 cm −1 where the hydrogen‐out‐of‐plane modes can be detected may indicate that the binding of imidazole does not increase the level of distortion of the haem plan with respect to the unbound protein.…”

“…In particular, the measurement in water‐based solutions at room temperature gives us the possibility to observe the structural deformations of the cofactor in its nearly native conformation or its dynamic evolution when perturbed (e.g., upon haem‐ligand binding). Consequently, transient species between different active conformations of the globin can be observed as well, giving a more dynamic picture of the molecule of interest when compared with crystallography . The resonance‐enhanced ROA spectra exhibit good S/N ratio, despite the use of much lower amount of sample with respect to the far‐from‐resonance conditions.…”

Chiral Amplification in Nature: Studying Cell‐Extracted Chiral Carotenoid Microcrystals via the Resonance Raman Optical Activity of Model Systems

“…Being structurally sensitive,t he HOOP band [37,38] can be considered as evidence for the presence of deuterated (more bent) bCm olecules in the supramolecular mixed arrangement, which gives achiral response.Moreover,anincrease in the intensity of the HOOP band can also be seen in the Raman spectra of the bCmonomer and its aggregate before, as well as after deuteration ( Figure S5). However,b ased on the vibrational analysis of 11,12-dideuterioretinal, [40] it can be seen that the HOOP band is enhanced for deuterated derivatives,w hich confirms as ubstantial deuterium-induced change in the molecular structure of bC-d compared with bC.…”

“…[9] In contrast to bC, amodel system built from the combination of achiral bC(80 %) and chiral aC(20 %) shows ac hiral signal in the ECD and RROA spectra (Figure 2, right). [37,38] Therefore,i ts occurrence is most likely related to the bending of the carotenoid chain in the bC/ aCmodel, which probably comes from aC, whose structure is more bent ( Figure S2). [37,38] Therefore,i ts occurrence is most likely related to the bending of the carotenoid chain in the bC/ aCmodel, which probably comes from aC, whose structure is more bent ( Figure S2).…”