Researches On Characteristics Of Phycoerythrins And Phycobilisomes From Polysiphonia Urceolata

Red algae are important group in algae. They have important roles in researches on marine ecology and biological evolution and can be used as foods and drugs. Specificity of red algae is the light-harvesting antenna system, phycobilisomes which are assembled with phycobiliproteins. Phycobilisomes of red algae are derived from cyanobacteria, but are much more complex than those of cyanobacteria. At present, researches on phycobiliproteins and phycobilisomes (especially phycobilisomes) are mainly concentrated in cyanobacteria, and researches on red algae are mainly on the lower red algae, e.g. Porphyridium and Porphyra. Researches on structure of phycobilisomes from red algae are few. Characteristics of phycoerythrin and phycobilisomes from a marine red algae Polysiphonia urceolata are studied in this paper to provide some foundation for clarifying the structure of phycobilisomes from the red algae.Proper conditions of SDS-PAGE for subunit analysis of R-phycoerythrin are studied in this paper, and proper conditions under which the subunits can be well separated have been found. The critical factors affecting protein SDS-PAGE are SDS concentration and ionic strength, and the mechanisms should be associated with critical micelle concentration (cmc) of SDS and ratio of SDS monomer to micelle in SDS-PAGE system. This study may also provide some references for subunits analysis of other proteins.Phycoerythrins from Polysiphonia urceolata are composed of four types of subunits:α,β,γ1 and γ2. Two bands whose molecular weights are about 40kDa and 50kDa are found in SDS-PAGE gels of the phycoerythrins. Studying the composition of these two bands and analyzing the cause of their formation can provide some basis and references on clarifying the position of the chromophores, especially those of γ subunit, and studying the relationships between the subunits of phycoerythrin.Phycoerythrins extracted with different methods vary much when they are analyzed by gel filtration chromatography column of Sephadex G-150. Trimeric phycoerythrin having no y subunit is found by analyzing the aggregate types of crude phycobiliprotein prepare by different methods, and trimeric phycoerythrin are tend to be dissociated into monomers.In research on phycobiliproteins from dissociated phycobilisomes, three types of phycoerythrins have been found:two types of phycoerythrins having different y subunit and one type phycoerythrins having no y subunit.Fluorescence characteristics of phycobilisomes are studied in this research, and possible energy transfer paths are proposed by analyzing the fluorescence spetra of phycobilisomes and phycoerythrins.The phycobilisomes are relatively stable in the high concentrations of phosphate buffers (PBS), but tend to be dissociated in the low concentration of PBS. The components of phycobilisomes treated with 500mmol/LPBS were analyzed. It shows that Reduce the concentration of the phosphate buffer, phycoerythrins are more easily dissociatd from phycobilisomes when the concentration of PBS is reduced; and phycoerythrins are tend to be dissociated a whole "rod" once. These all prove that the "rods" in structures of phycobilisomes from red algae are not independent like those in cyanobacteria but are stacked together.Different solutions have different effects on stability of phycobilisomes: monovalent cations have better effects than divalent cations, and the larger the molecule, the better the effects; sulfates and phosphates have good effects; as for monovalent anion, chlorides and nitrates have poor effects, but acetates have better effects. Ions should affect the stability of phycobilisome by screening the charges on surfaces of phycobiliproteins in phycobilisome; thereby repulsions between phycobiliproteins are weakened. The strength of repulsions between the phycobiliproteins is the essential factor influencing the structural stability phycobilisomes.Phycoerythrin-phycocyanin complexes have been obtained in preparation phycobilisomes. Chromophores in complex which have higher absorption at 587nm have been found by analyzing the absorption difference spectra of complexes and phycoerythrins, but their absorption characteristics change when they are dissociated. Chromophores having higher absorption at 587nm are also found in the dissociation products of phycobilisomes in 200mmol/L PBS, and their fluorescence emission peak are at about 604 nm. Discovery of 587nm chromophore can make it easy to explain the efficient energy transfer between phycoerythrins and phycocyanins:the 587nm chromophores accept and centralize light energy transferred by other phycoerythrin chromophores, and transfer the energy to the phycocyanin PCB chromophore efficiently. The phycoerythrins having γ2 subunit are believed to be closer to the "core".Phycoerythrin and phycobilisomes were cross-linked using formaldehyde separately. There are two α subunits crosslinked together in phycoerythrin after treatment of formaldehyde; and a α subunit can be linked to a β subunit in phycobilisomes treated with formaldehyde; the γ subunits are all involved in cross-linking after treatment both in phycoerythrin and phycobilisomes.Phycoerythrins and phycobilisomes were treated two kinds of proteases. The phycoerythrin hexamer will not be dissociated after treatments of proteases, but the β subunits will be cut into 14.4kDa peptides; there are only few allophycocyanins after phycobilisomes are digested by proteinase K. "Rods" and phycoerythrin-phycocyanin complex have been isolated after phycobilisomes are digested by proteinase K. It shows from the absorption characteristics of phycobilisomes isolated from digestion products phycobilisomes that the structures of "cores" in phycobilisomes of red algae may be different with cyanobacteria. The discovery of the "rods" can supply some foundation to researches on structure of phycobilisomes from red algae.