Abstract
Phycobilisomes (PBS) are accessory light harvesting protein complexes formed mainly by phycobiliproteins (PBPs). The PBPs absorb light that is efficiently transferred to Photosystems due to chromophores covalently bound to specific cysteine residues. Besides phycobiliproteins (PE), the PBS contains linker proteins responsible for assembly and stabilization of the whole complex and the tuning of energy transfer steps between chromophores. The linker (γ33) from Gracilaria chilensis, is a chromophorylated rod linker associated to (αβ)6 hexamers of R-phycoerythrin (R-PE). Its role in the energy transfer process is not clear yet. Structural studies as well as the composition and location of the chromophores are essential to understand their involvement in the energy transfer process in PBS. To achieve this, the coding gene of γ33 was cloned and sequenced. The sequence was analyzed by informatics tools, to obtain preliminary information which leaded the next experiments. The protein was purified from R-phycoerythrin, and the sequence confirmed by mass spectrometry. The coding sequence analysis revealed a protein of 318 aminoacid residues containing a chloroplastidial transit peptide (cTP) of 39 aminoacids at the N-terminus. The conservation of cysteines revealed possible chromophorylation sites. Using α and β R-PE subunits as spectroscopic probes in denaturation assays, we deduced a double bonded phycourobilin (PUB) on γ33 subunit that were confirmed between Cys62 and Cys73 (DL-PUB62/73) by mass spectrometry. The cysteines involved in the double link are located in a helical region, in a conformation that reminds the position of the DL-PUB50/61 in the β subunit of R-PE. The position of single linked PUB at Cys95 and a single linked PEB at Cys172 were also confirmed. Spectroscopic studies show the presence of both types of chromophores and that there are not energy transfer by FRET among them.
Highlights
Phycobilisomes (PBS) are highly efficient accessory light harvesting protein megacomplexes, responsible for the conduction of energy towards the photosynthetic reaction centers present in cyanobacteria and red algae
This report presents the position in the sequence and identity of the chromophores, information that, with the structural data provided by Zhang et al, (2017) [12] would allow us to advance in the elucidation of functional aspects of γ subunits on our energy transfer model in PBS
The γ33 subunit of R-PE is a protein of 279 residues organized in 2 internal repeats that suggest its arrangement as domains
Summary
Phycobilisomes (PBS) are highly efficient accessory light harvesting protein megacomplexes, responsible for the conduction of energy towards the photosynthetic reaction centers present in cyanobacteria and red algae They absorb light in a wavelength range where chlorophyll is poorly efficient (the green gap), and transfer energy directionally to the photosystem II [1]. It has been proposed that linkers are positioned in the central cavity of PBP trimers or hexamers [13] This location would disrupt the symmetry of the possible energy transfer pathways between chromophores of the complexes, guiding and promoting the energy transfer along the rods and core [14]. This report presents the position in the sequence and identity of the chromophores, information that, with the structural data provided by Zhang et al, (2017) [12] would allow us to advance in the elucidation of functional aspects of γ subunits on our energy transfer model in PBS
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