Crystal Structure Of The EsaG/Orf13/EsaH Complex From The Type ? Secretion System In Edwardsiella Tarda

Edwardsiella tarda(E.tarda)is an important wide host-range gram-negative pathogenic bacteria,which can infect fish,amphibians,reptiles,mammals and even humans.Fish,both marine and freshwater fish,has been the most common host of E.tarda.In aquaculture industry,E.tarda is one of the most common pathogenic bacteria.Edwardsiellosis caused by E.tarda has led to huge economic losses every year.Although several groups are focusing on vaccine development,commercial vaccines are not yet available.Type III Secretion System(T3SS)is widespread among gram-negative bacteria pathogenic for animal and plants.Pathogens use T3SS to "inject" virulence proteins(effectors)into the cytoplasm of host cells to modify host cell physiology.T3SS,is essential for its bacterial pathogenesis by delivering effectors into fish cells.Fish which infected E.tarda may suffer from septicemia.The needle of T3SS is made up of more than 100 needle protein subunits.Studies show that the needle of T3SS is oligomerized by the C terminus of needle protein through hydrophobic interactions.EsaG is the needle subunit of E.tarda T3SS.Although needle proteins show low sequence homology,they possess conserved 3D structures.Domain analysis suggests that Orf13 belongs to YscE superfamily.Since YscE is one of the two chaperone protein of Yersinia needle protein we presume that Orf13 is the chaperone of E.tarda needle protein.Moreover,yeast two hybrid assay showsthat EsaH have stronge interaction with Orf13.This result suggests that EsaH may server as the other chaperone of EsaG.Here we solved crystal structures of Orfl 3/EsaH and EsaG/Orfl 3/EsaH complex by crystallography.The atomic structure of EsaG/Orf13/EsaH elucidates the mechanism how Orf13/EsaH complex protects the needle protein,EsaG.The crystal structure reveals that the C terminus of the needle protein EsaG is engulfed within the hydrophobic groove of the Orf13/EsaH while the N terminus of that is disordered free from the Orf13/EsaH complex.Our results suggest that Orf13 and EsaH are required for the stabilization of EsaG so as to prevent its oligomerization.Through sequence and structure alignment,we found that Orf13/EsaH complex can still act as needle chaperone although EsaH possesses only 5? helices,suggesting that E.tarda is more conducive to survive from an evolution point of view.To our knowledge,this is the first structure of E.tarda T3SS and we figured out the mechanism how Orf13/EsaH protects the needle protein.Due to the importance of needle protein for bacterial pathogenesis,the EsaG/Orf13/EsaH may serve as the target of antibacterial drug and the complex structure may provide basis for drug design and vaccine development.