The Structure Biology Research Of Synbindin And P97-CTD Of EIF4G Family

Transport protein particle(TRAPP) is a large multiprotein complex involved in EG-to-Golgi and intra-Golgi traffic.It is found in both yeast and human and involves in the progress that tethering the vesicles to their distinct membrane.The BET3p, BET5p,Trs20p,Trs23p,Trs31p,Trs33p,Trs85p is common subunits in TRAPPⅠand TRAPPⅡ,and Trs65p,Trs120p,Trs 130p is additional subunits of TRAPPⅡ.These two forms of TRAPP act at different stages of membrane traffic.TRAPPⅠbind endoplasmic-reticulum derived vesicles to Golgi while TRAPPⅡis involved in transport within the Golgi.Synbindin,the human orthologue of yeast Trs23p,was first identified by a yeast two-hybrid screening using the syndecan-2 cytoplasmic domain as bait.In hippocampal neurons,synbindin was suggested that the synbindin/syndecan complex is involved in synaptic membrane trafficking and thereby regulates the formation of dendritic spines.We report the crystal structure of Synbindin,determined by the Se-MAD method at 2.8A resolution.Due to the flexibility of the atypical PDZ domain,we get the NMR structure of this domain.The crystal structure of Synbindin consists by two domains:the APD domain at N-terminal which spited into the Longin domain into two part.The synbindin LD represent the first split LD structure among known Longin domains.This Longin domain contain five-stranded antiparallelβ-sheet at the center sandwitched by one helix(α1_L) in one side and two helices(α2_L and a 3_L) in the other side.And the APD domain has something different with the typical PDZ domain.Using the SPR, ITC and NMR titration experiment,we find the interaction between the cytoplasmic domain of Syndean-2 and APD is quit weak.Analysis the crystal structure of Synbindin and compared with structure of other components of TRAPP complex,we suggest the possible interaction model of longin domain.Eukaryotic translation initiation is a sophisticated cellular process,which is stimulated by a number of proteins called eukaryotic initiation factors(elFs).eIF4G acts as a scaffold protein.It can bind to eIF4A and elF4E to form a large complex eIF4F.It can also recruit eIF3,which in turn binds the small ribosomal subunit. eIF4G also binds the poly(A) binding protein(PABP),the Mnkl protein kinase,which phosphorylates eIF4E.p97(also known as DAPS) is a modulator of translation initiation with homology to the middle and the C-terminal regions of eIF4G.It may function like a dominant negative inhibitor of translation initiation by acting as a molecular mimic of eIF4G.We solve the crystal structure of C-terminal domain of p97,determined by the Se-MAD method at 2.0A resolution.We discover that the crystal structure of C-terminal domain of p97 contains two atypical HEAT domains at C-terminal,which is structurally similar to C-terminal of eIF4G,although the two protein share very little amino acid sequence similarity.