Clonging And Bioinformatics Analysis Of T6SS Chaperone Protein ClpV From Aeromonas Hydrophila

Aeromonas hydrophila is an important pathogen that causes bacterial hemorrhagic disease on a variety of freshwater fish.It becomes one of the important factors which restrict the development of freshwater aquaculture.So the study on the pathogenic mechanism of the bacteria which has practical significance to prevent and control of the infection,occurrence and spread of the disease caused by the the bacteria.The VI type secretory system(T6SS)is a new type of secretory system which is widely distributed in recent years and is closely related to the pathogenicity of bacteria.At present,there has been some progress in the research of A.hydrophila T6 SS.However,the interaction of the whole secretion system elements,signal transduction pathway and regulation mode is still not clear.This paper makes preliminary research on the function of ClpV using bioinformatics to systematic analyse the characteristics of sequence,the signaling pathway,the interactional network of protein associated with ClpV and GO annotation.Those results maybe proved a certain foundation on the further exploring the mechanisms of regulated and pathogenic of ClpV.Obtained the full length of cDNA sequence of ClpV,and analyzed the sequence alignment and phylogenetic tree.The results showed that the gene sequence had a complete open reading frame(ORF)in a full length of 2643 bp.Analysis found that the similarity of homology with related proteins different from other species were 81-93%,and Aeromonas enterobiosis is the highest homology(93%).ClpV encodes a protein of 880 amino acid residues with the molecular weight of 96748.39 and the isoelectric point of p I = 5.23.Using the on-line analysis tools and software of bioinformatics to predicte and analyze the characteristics of ClpV gene including the composition of amino acid,physicochemical properties,hydrophobicity/hydrophilicity,signal peptide,conserved domain,analysis of homology and other aspects.The predicted and analysis results showed that ClpV was the hydrophilic protein without signal peptide,transmembrane structure,and GPI modification sites.Meanwhile,ClpV also contain multiple phosphorylation sites and glycosylation sites.The analytical prediction of the primary structure,the secondary strcture and the tertiary structure of ClpV was done.Acquired the characterics of the primary structure and the functional domain of ClpV.The secondary strcture include alpha helix,beta fold,beta rotation and random curl,which are mainly composed of alpha helix and random coil.The tertiary structure was automatically searched,modeled and obtained the format of PDB results.The signal pathway of ClpV was predicted by KEGG software of bioinformatics,and obtained the map of signal pathway of ClpV.The results indicate that ClpV was located on the inner membrance of VI secretion system,and was adjacent to VgrG,‘tubulose'-like Hcp,Icm F,DotU and Fha1.Using the STRING database to search for the interactional network of protein associated with ClpV.Get 10 most reliable protein and protein interaction network diagrams.Using http://www.geneontology.org/ to search for the Gene Ontology annotation of ClpV and the results show that the protein involved in as many as 26 kinds of biological processes.The majority of which involved in the metabolism process,such as cytokine metabolic process,Proteolysis,biosynthetic process,positive regulation of protein metabolic process,negative regulation of protein metabolic process,insulin metabolic process,receptor biosynthetic process.The molecular function of ClpV play mainly in the form of complex.In addition,there is no localization of ClpV about cell components which is consistent with the results of prediction before,the ClpV cells located in the cytoplasm.