Phycoerythrin
Phycoerythrin is a red protein-pigment complex from the light-harvesting phycobiliprotein family, present in red algae and cryptophytes, accessory to the main chlorophyll pigments responsible for photosynthesis.
Like all phycobiliproteins, it is composed of a protein part covalently binding chromophores called phycobilins. In the phycoerythrin family, the most known phycobilins are: phycoerythrobilin, the typical phycoerythrin acceptor chromophore, and sometimes phycourobilin. Phycoerythrins are composed of monomers, usually organised in a disk-shaped trimer 3 or hexamer 6. These typical complexes also contain a third type of subunit, the γ chain.
Phycobilisomes
Phycobiliproteins are part of huge light harvesting antennae protein complexes called phycobilisomes. In red algae they are anchored to the stromal side of thylakoid membranes of chloroplasts, whereas in cryptophytes phycobilisomes are reduced and are densely packed inside the lumen of thylakoides.Phycoerythrin is an accessory pigment to the main chlorophyll pigments responsible for photosynthesis. The light energy is captured by phycoerythrin and is then passed on to the reaction centre chlorophyll pair, most of the time via the phycobiliproteins phycocyanin and allophycocyanin.
Structural characteristics
Phycoerythrins except phycoerythrin 545 are composed of monomers assembled into disc-shaped 6 hexamers or 3 trimers with 32 or 3 symmetry and enclosing central channel. In phycobilisomes each trimer or hexamer contains at least one linker protein located in central channel. B-phycoerythrin and R-phycoerythrin from red algae in addition to α and β chains have a third, γ subunit contributing both linker and light-harvesting functions, because it bears chromophores.Porphyridium cruentum. The asymmetric unit 2 on the left and assumed biological molecule 3. It contains, and.
R-phycoerythrin is predominantly produced by red algae. The protein is made up of at least three different subunits and varies according to the species of algae that produces it. The subunit structure of the most common R-PE is 6γ. The α subunit has two phycoerythrobilins, the β subunit has 2 or 3 PEBs and one phycourobilin, while the different gamma subunits are reported to have 3 PEB and 2 PUB or 1 or 2 PEB and 1 PUB. The molecular weight of R-PE is 250,000 Daltons.
Crystal structures available in the Protein Data Bank contain in one 2 or 2 asymmetric unit of different phycoerythrins:
of chlorophyll for example, but tetrapyrrole is linear, not closed into ring with metal ion in the middle.
Gracilaria contains R-phycoerythrin.
cryptophyte Rhodomonas CS24. Colors: chains -,,, ,, ,,,,.
The assumed biological molecule of phycoerythrin 545 is 2 or rather. The numbers 2 and 3 after the α letters in second formula are part of chain names here, not their counts. The synonym cryptophytan name of α3 chain is α1 chain.
The largest assembly of B-phycoerythrin is 3 trimer. However, preparations from red algae yield also 6 hexamer. In case of R-phycoerythrin the largest assumed biological molecule here is 6, or 6 dependently on publication, for other phycoerythrin types 6. These γ chains from the Protein Data Bank are very small and consist only of three or six recognizable amino acids, whereas described at the beginning of this section linker γ chain is large . This is because the electron density of the gamma-polypeptide is mostly averaged out by its threefold crystallographic symmetry and only a few amino acids can be modeled.
For 6, 6 or the values from the table should be simply multiplied by 3, 3 contain intermediate numbers of non-protein molecules.
In phycoerythrin PE545 above, one α chain binds one molecule of billin, in other examples it binds two molecules. The β chain always binds to three molecules. The small γ chain binds to none.
Two molecules of N-methyl asparagine are bound to the β chain, one 5-hydroxylysine to α, one Mg2+ to α-3 and β, one Cl− to β, 1-2 molecules of SO42− to α or β.
Below is sample crystal structure of R-phycoerythrin from Protein Data Bank:
Gracilaria chilensis - basic oligomer 2. It contains,,,,. One fragment of γ chain is red, second one white because it is not considered as alpha helix despite identical aminoacid sequence.
of R-phycoerythrin from Gracilaria chilensis 6 .
Spectral characteristics
s in the visible light spectrum are measured at 495 and 545/566 nm, depending on the chromophores bound and the considered organism. A strong emission peak exists at 575 ± 10 nm.| Property | Value |
| Absorption maximum | 565 nm |
| Additional Absorption peak | 498 nm |
| Emission maximum | 573 nm |
| Extinction Coefficient | 1.96 x 106 M−1cm−1 |
| Quantum Yield | 0.84 |
| Brightness | 1.65 x 106 M−1cm−1 |
PEB and DBV bilins in PE545 absorb in the green spectral region too, with maxima at 545 and 569 nm respectively. The fluorescence emission maximum is at 580 nm.
R-Phycoerythrin variations
As mentioned above, phycoerythrin can be found in a variety of algal species. As such, there can be variation in the efficiency of absorbance and emission of light required for facilitation of photosynthesis. This could be a result of the depth in the water column that a specific alga typically resides and a consequent need for greater or less efficiency of the accessory pigments.
With advances in imaging and detection technology which can avoid rapid photobleaching, protein fluorophores have become a viable and powerful tool for researchers in fields such as microscopy, microarray analysis and Western blotting. In light of this, it may be beneficial for researchers to screen these variable R-phycoerythrins to determine which one is most appropriate for their particular application. Even a small increase in fluorescent efficiency could reduce background noise and lower the rate of false-negative results.
Practical applications
R-Phycoerythrin is useful in the laboratory as a fluorescence-based indicator for the presence of cyanobacteria and for labeling antibodies, most often for flow cytometry. Its use is limited in immunofluorescence microscopy due to its rapid photobleaching characteristics. There are also other types of phycoerythrins, such as B-Phycoerythrin, which have slightly different spectral properties. B-Phycoerythrin absorbs strongly at about 545 nm and emits strongly at 572 nm instead and could be better suited for some instruments. B-Phycoerythrin may also be less "sticky" than R-Phycoerythrin and contributes less to background signal due to non-specific binding in certain applications. However, R-PE is much more commonly available as an antibody conjugate.R-Phycoerythrin and B-Phycoerythrin are among the brightest fluorescent dyes ever identified.