Research On High-efficiency Energy Transfer Mechanism Of Cyanobacterial Phycobilisomes-Synechocystis Sp.PCC 6803 And Thermosynechococcus Vulcanus NIES 2134

Phycobilisomes(PBSs) are water-soluble light-harvesting pigment complexes found in red algae and cyanobacteria.PBSs can transfer energy to the reaction center(RC)with an efficiency of more than 95%,and this efficiency has always attracted attention.In recent years,with the maturity of ultrafast laser spectroscopy technology,the study of the photosynthesis energy transfer between internal atoms and molecules of phycobiliproteins and photosystem II(PSII)has increasingly become a hot issue in the interdisciplinary research of physics and life sciences.Studying the energy transfer mechanism of PBSs can provide a theoretical basis for the development of underwater blue-green light-harvesting devices or artificial photosynthetic devices.Therefore,this experiment used physiological and biochemical technology combined with spectroscopy to study the multi-scale energy transfer process of PBSs at the supermolecular complex level(From Synechocystis sp.PCC 6803 and Thermosynechococcus vulcanus NIES 2134,referred to as S.6803 and T.2134 respectively.),component level(recombinant core structural protein of PBSs-r APC)and cell level.Based on the research results,the relationship between the structure and energy transfer of PBSs were analyzed.The main contents are as follows:(1)In order to reveal the energy transfer dynamics of PBSs,we used S.6803 PBSs as the material,and analyzed the structural assembly and spectral characteristics of S.6803 PBSs by measuring its steady-state absorption and fluorescence emission spectra.The results showed that the main absorption peak at 621 nm of S.6803 PBSs was derived from the phycocyanin(PC)chromophores,and this absorption peak was significantly red-shifted compared with PC that was not assembled into PBSs,which reflected the complexity of chromophore microenvironment in PBSs.Through 77 K time-resolved fluorescence emission spectroscopy,the energy transfer process of PBSs was studied,and the results showed that there are three time components in the energy transfer process of PBSs: 9 ps,115 ps and 1680 ps,where 9ps represented the energy transfer time from PCs to APCs,115 ps represented the energy transfer time from the PBS rod to the PBS core,and 1680 ps represented the time from when the energy was absorbed to when it was emitted as fluorescence from the terminal energy acceptor of the S.6803 PBSs.This result confirmed that there was a multi-time scale energy transfer process in PBSs;CD spectroscopy results show that there are two pairs of pigment exciton pairs in the PBS core.Except for the exciton pair in APCs at 660 nm,there also be exciton pairs in the terminal energy acceptors at 670 nm.The existence of exciton pairs may be the reason for the high-efficiency energy transfer in the PBS core and the multi-time-scale energy transfer in PBSs.Since the three-core six-rod PBS possessed by S.6803 is widely present in cyanobacteria,the results of this study provide a data reference basis for understanding the high-efficiency energy transfer process of cyanobacterial PBSs.(2)In order to reveal the relationship between the energy transfer process of PBSs and their structures,we chose T.2134 PBSs that has a special rod-core connection component as the experimental material,and studied their energy transfer kinetics,energy transfer pathways and related transfer rates.The results showed that in T.2134,the energy transfer time from PCs to APCs was 10 ps,the energy transfer time from the PBS rod to the PBS core was 80 ps,and the time of energy transfer from absorption to emitted as fluorescence from the terminal energy acceptors is 1250 ps.Comparing the energy transfer process of S.6803 PBSs,it can be found that,in order to adapt to the high temperature environment,the structure of T.2134 PBSs is different from that of S.6803 PBSs which results in a different transfer time between them.This further indicates that the structural difference of PBSs will affect the energy transfer process of PBSs.CD spectroscopy results showed that there were two pairs of pigment exciton pairs in the core structure of PBSs,located in the APC and the energy terminal receptor proteins,respectively.It shows that T.2134 PBSs will not only evolve a more stable PBS structure in order to cope with extreme environments,but also the tightly arranged and strongly coupled exciton interactions of phycobilins can also ensure the efficiency of energy transfer.(3)In order to reveal the efficient energy transfer mechanism of the PBS core,the genetically recombined allophycocyanin-r APC(from S.6803)was selected as the material.The energy transfer kinetics of r APC was studied by measuring the steadystate spectra,CD spectra and various time-resolved spectra.The results showed that r APC and natural APC could maintain the same spectral characteristics and active conformation under the test conditions;And in r APC trimer,the ?84PCB and ?84PCB can form excitonic pigment pairs,which is believed to improve the energy transfer efficiency of the trimer through exciton splitting;In addition,the energy transfer process of r APC trimer was studied through ultrafast time-resolved spectroscopy.It was found that the energy transfer time from 620 nm to 650 nm is 300~600 fs and the decoherence time may occur within 19 fs;Further detection by two-dimensional electronic spectroscopy and broadband transient grating found that the coherent oscillation frequencies in the range of 660 cm-1 and 820 cm-1 in r APC trimer was considered to be most likely to be related to the coherent energy transfer process of vibration coupling and may promote the energy transfer.Since the superposition state of coherent state energy transfer has the characteristics of quantum computing,the optimal path can be given through a single quantum calculation,and energy can be efficiently and accurately transferred to the target molecule through one energy transfer step.Therefore,the above results provide a data basis for the study of the efficient energy transfer mechanism of the cyanobacterial PBS core structure.(4)In order to reveal the influence of changes in PSII photosynthetic electron transfer rate on the structure of PBSs,S.6803 cells were chosen as the experimental material.The growth of S.6803 was monitored and chlorophyll fluorescence detection was carried out.It was found that the growth and the photosynthetic electron transfer rate of S.6803 showed a Cu-Fe concentration association.Cell steady-state spectroscopy results showed that the content of PBSs in cells was affected by the CuFe concentration association,which showed that the content ratio of PBSs to chlorophyll protein was affected by electron transfer rate under the condition of sufficient PBS synthetic elements(the synthesis of phycocyanins was not restricted).However,the change of copper concentration alone had a relatively weak effect on the content of PBSs.The extraction,structure identification and steady-state spectroscopy results of PBSs showed that changes in iron concentration would affect the synthesis of PBSs,but would not affect the structure stability of the intact PBSs.The change of copper concentration would change the structural stability of PBSs,and the change trend of stability was consistent with the electron transfer rate associated with copper concentration.We think that the change in structural stability may be due to the structural change of PBS rods.These results all confirmed that the change of photosynthetic electron transfer rate would affect the structure of PBSs.This effect may be acting on the rod structure of PBSs to regulate the light-harvesting efficiency of PBSs.Based on the energy transfer mechanism results of the multi-scale PBSs samples,the following conclusions can be drawn: there are two time-scale energy transfer processes of intact PBS complexes and two time-scale energy transfer processes in the recombinant PBS core structural protein-r APC,which proves that there are both fast and slow energy transfer mechanisms during the energy transfer process of PBSs.Pigment exciton pairs exist in the core structure of PBSs,and pigment exciton pairs are also existing in the recombinant PBS core structural protein-r APC.The existence of pigment exciton pairs is likely to be related to the rapid energy transfer process of PBSs,and may trigger the coherent resonance energy transfer process.Moreover,the structure evolution of PBSs is affected by the algal habitat,and the structure determines their energy transfer process.In addition to the external environment affecting the structure of the PBSs,changes in the internal photosynthetic electron transfer rate will also affect the structure of PBSs,to optimize the energy transfer efficiency.