Study On The Function,structure Of AP-B And ApcE-related Non-photochemical Quenching,state Transition Of Synechocystis Sp. PCC 6803

Cyanobacteria has a unique antenna complex,the phycobilisome(PBS),they harvest a significant portion of the solar energy that is globally used by natural photosynthesis.Allophycocyanin B(AP-B)and core-membrane linke(LCM)are the two terminal emitters in phycobilisomes,the unique light-harvesting complexes of cyanobacteria and red algae.Their low excitation energy level and the correspondingly red-shifted absorption and fluorescence emission play an important role in funneling excitation energy from the hundreds of chromophores of the extra-membraneous phycobilisome to the reaction centers within the photosynthetic membrane.In PBS,both AP-B and core-membrane linker(LCM)are in very low abundance,the molecular basis for the extreme red-shift and directional energy transfer is largely unknown.In this work,we firstly obtained the apc D fragment which contains the His-tag sequence in carboxyl terminal and then constructed the ApcD-His mutant.AP-B carrying a His tag was purified from Synechocystis sp.PCC 6803 and crystalized by hanging-drop vapour-diffusion method.Finally,we obtained the crystal structure of trimeric AP-B(ApcD-ApcB)3 at 1.75 ?.The red-shift of the chromophore results from interactions with both subunits that render the conjugation system coplanar.We propose that the enhanced coplanarity effectively extends the conjugation system of PCB and leads to the red-shifted absorption(669 nm)and fluorescence emission(679 nm)of the ApcD chromophore in AP-B,thereby enabling highly efficient energy transfer from the phycobilisome core to the reaction centers.Owing to the high symmetry in space group I432,the study of the crystal structure of trimeric AP-B would inspire us to design biomolecular devices based on phycobiliproteins.Phycobilisomes are consisting of phycobiliproteins,serving mainly for photosystems ?.Cyanobacteria as a kind of photosynthetic organisms,has a lot of light protection mechanisms.There are mainly state transition and non-photochemical quenching(NPQ)which induced by blue high light.On the state transition,most scholars all agree with this idea: the phycobilisomes can move on the thylakoid membrane,so adapting to the low light variation.But now the detailed mechanism of state transition is still not known.On NPQ that induced by blue high light,the present study shows that NPQ is the activated orange carotenoid protein quenches the excitation energy in phycobilisomes through non-photochemical quenching under blue high light,so photoprotecting photosystem.But the specific binding sites of OCP with PBS are still unknow.Owing to apc E connecting phycobilisome and thylakoid membrane simultaneously,not only participates in the assembly of PBS but also determines its size.We according to the characteristics of structure and function in apcE,by using the method of insertional inactivation,we truncated away the third linker repeat(Rep3),the second arm(Arm2),both the second arm(Arm2)and third linker repeat(Rep3)from the phycobilisome core-membrane linker(ApcE)respectively,and we obtained the ?(rep3),?(arm2)and ?(arm2/rep3)mutants.Finally,we analyzed the physiological activity,spectral properties,NPQ and state transition of these mutants.In this Synechocystis ?(rep3)mutant,both the state transition and non-photochemical quenching were well maintained.After the second arm(Arm2),both the second arm(Arm2)and third linker repeat(Rep3)were truncated away from ApcE,the non-photochemical quenching was maintained but the state transition disappeared in the ?(arm2)and ?(arm2/rep3)mutants.Therefore,it indicates that non-photochemical quenching resides at the two cylinders in the basal layer and the state transition relates to the Arm2.Our results could contribute to study the light protection mechanism in cyanobacteria.