Exploring Physiological Roles Of PSB27 In Arabidopsis

In Cyanobacteria,PSB27 has a lipobox domain that can reaction with membrance.The upsteam of this domain has many hydrophobic animo acids,which help protein binding to membrance.PSB27 is involved assembly and repair of the PSII complex thought linking with loop E of CP43.Under experiment conditions,manganese cluster binding to PSII to oxidize water in the help of OEC.The mutant showed no difference with wild type.Under abiotic stresses especially high light,Psb27 binds to PSII without OEC and regulate assembly of the Mn4CaO5 cluster during PSII repair.While in the Arabidopsis,PSB27 located in the lumen.Under experiment conditions,mutant show same phenotype with wild type;while under high light,The mutant show lower PSII efficiency,less starch,less anthocyanin but more ROS.Even PSB27 is not essential for living but its mutant showed extremely sensitive to light.We suggested that PSB27 protein play an important role in the pathway of light protection.But the mechanism of PSB27 has remained elusive.We want to know who linked with PSB27 and how does it work in the light protection and photo inhibition in Arabidopsis.The first step is to get Psb27 antibody from rabbit,we expressed PSB27 mature protein in Ecoil,then injure protein into rabbit,Psb27 antibody would get from rabbit's blood.Subsequently,we constructed plasmid pB003 with Psb27-His or Psb27-2HA,and transfered them into psb27 mutant in the help of Agrobacterium GV3101 to obtain the transgenic plants.Used PSB27 antibody do western bloting to check the location of PSB27 and the protein levels in transgenic plants.Because Histidine can bind with nickel,and Histidine was coexpressed with PSB27,so we can use nickel to get PSB27 and the reaction proteins,then analysis the reaction proteins by mass spectrometry.PLDALPH1 encodes phospholipase D alpha 1,it has GTPase activator activity,bind with calcium ion and protein.LHCB1.3 is chlorophyll A-B binding protein.FTSH1 bind with Protein,metal ion and ATP,it has ATP-dependent peptidase activity and ATPase activity,involve in photoinhibition,photosystem II repair.NUA has nucleocytoplasmic transporter activity.PSBS binds to chlorophyll and xanthophyll binding,and has substrate-specific transmembrane transporter activity.TIC62 has transmembrane transporter activity.FTSH8 binds to zinc and ATP,and it has ATP-dependent peptidase activity,ATPase activity.P40 and CP31A bind to RNA.PNSL1 binds to calcium ion,and involve to photosynthesis.Legume Lectin binds to carbohydrate binding,and has kinase activity.As we all know,The core of PSB27 protein is a right-handed four-helix bundle and linked by five loops.In order to know which animo acids or loops are important points to the stabily of PSB27 structure.Take 15 kinds of polar amino acids and 5 loops into alanine which has small size and simple structure.When screened those transgenic plants in protein levels by western bloting,we found multimutantion likeD3E4E5K8D9 and K86R87K88 show very less expression of mature protein.So we guess those points may effect that PSB27 entered into the lumen.Finally we take psb27 gene from Cyanobacterium Synechocystis sp.PCC7942 and Chlamydomonas Reinhardtii into mutant in Arabidopsis to detected if PSB27 has same fountion in different species.