Photophysics of phycoerythrocyanins from the cyanobacterium Westiellopsis prolifica studied by time-resolved fluorescence and coherent anti-Stokes Raman scattering spectroscopy

Abstract

Three building blocks of the antenna complexes of the cyanobacterium Westiellopsis prolifica were studied: PEC(X), which is similar to the ot-subunit of phycoerythrocyanin (PEC), trimers of PEC and monomers derived from these by deaggregation with KSCN. The fit of the fluorescence decay curve of PEC(X) requires at least four exponentials, although it supposedly contains only one chromophore. The coherent anti-Stokes Raman scattering (CARS) spectra indicate that the heterogeneity observed is due to geometrical isomers, which are in part generated by photoinduced processes. A similar heterogeneity in chromophore structure and properties is also found in the monomers, where four exponentials are needed to fit the fluorescence decay curve. As in trimers, there is a long-lived, low-amplitude component, which can be assigned to impurities and/or oxidation products. The energy transfer time between the two phyocyanobilin chromophores in the/3-subunit is about 500 ps; the lifetime of the fluorescing /3-chromophore is 1.5 ns. The phycoviolobilin chromophore in the a-subunit adopts different geometries characterized by fluorescence liftetimes of about 240 and 800 ps. No evidence was found for energy transfer between the otehromophore and the /3-chromophores. This energy transfer occurs in trimers on a time scale of less than 20 ps; the energy transfer time between the two different types of/3-chromophore is about 250 ps and the lifetime of the terminal emitter is about 1.5 ns. The excited state kinetics are therefore similar to those of PEC trimers from Mastigocladus laminosus, as are the CARS spectra, indicating a similar chromophore-protein arrangement. In comparison with phycocyanin, the ordering of the excited states of chromophores fl84 and/3155 may be changed. Although PEC trimers of IVestiellopsis prolifica show almost as good a photostability as trimers of Mastigocladus laminosus, monomers are so photolabile that no CARS spectra could be recorded.

Resonance‐Enhanced CARS Spectroscopy of Biliproteins: a Comparison Between Phycoerythrocyanin and Phycocyanin of Mastigocladus laminosus

“…It should also be mentioned that similar studies on the more photolabile PEC isolated from Wesriellopsis proli$cu (19) support the interpretation given above.…”

Fluorescence of Phytochrome Adducts with Synthetic Locked Chromophores

“…The phycoviolobilin chromophore of phycoerythrocyanin undergoes light-triggered Z to E isomerization around the C15 methine bridge. The fluorescence yield with the E-configured chromophore is very low (55).…”

Vibrational Spectroscopy of Phytochromes

“…Further techniques that circumvent fluorescence include coherent anti-Stokes (resonance) Raman spectroscopy (CARS) and surface-enhanced resonance Raman spectroscopy (SERRS). CARS probes the vibrational spectrum at the higher energy (anti-Stokes) side of the excitation line and, thus, does not interfere with fluorescence, but it is technically quite demanding [ 58 ]. SERRS can be applied when the target molecules are immobilized on nanostructured plasmonic metals like Ag or Au [ 59 , 60 ].…”