Study On Outer Membrane Protein Of Xanthomonas Axonopodis Pv.Citri

Citrus bacterial canker(CBC) disease, caused by Xanthomonas axonopodis pv.citri(Xac), is one of most serious diseases influence the production of citrus, and also serious quarantine diseases of citrus production.Xac is gram-negative bacterial,Outer membrane protein is one of an important component of gram-negative bacterial, which plays an important role in maintaining the bacteria normal morphology, physiology and biochemistry, metabolism and so on. And it has the immunogenicity and protective, playing a very important role in pathogenic bacteria infection processes. At present, the research of outer membrane protein of Xac is relatively less. In order to understand the nature, structure and function characteristics and to explore the function in the pathogenic process of outer membrane protein, this research study on them by means of bioinformatics analysis、cloning and expression. We got the information of the isoelectric point, molecular weight, signal peptide, domains and functions of Xac outer membrane protein by analysis of bioinformatics characters of Xac outer membrane protein. We choose typical outer membrane protein of Xac, XacOmp4、XacOmp5、 XacOmp7and XacOmp12, cloning them and expressing them in vetor of Pet28a and verifying the function of the XacOmp7.The results are concluded as follows:1. The protein of Xac has40outer membrane proteins. We divided these outer membrane proteins into20groups by ClustalX1.83according to there homology and detailed analyses their structure characteristics and functional structure domain by on line prediction. In addition to xacompl (1107aa) and xacomp2(2190aa), the other outer membrane proteins of Xac are between the size of the103aa-503aa,18groups with signal peptide,15groups of acidic proteins,2proteins with tranmembrane regions; In the outer membrane proteins of Xac, OmpW、OmpA、YadA and outer membrane efflux protein(OEP)are the main domains and account for65%of the total proteins. Further, the outer membrane proteins of Xac also have the structuralism module of SIP, TolueneX, Lipoprotein,SmpA-OmlA and others.2. Outer membrane proteins of Xac have five specific proteins by blastp with outer membrane proteins of Xanthomonas,15groups of outer membrane proteins of Xac have homologous protein in outer membrane proteins of Xanthomonas. Outer membrane proteins of Xac have the highest homology with Xcv. The types and numbers of the functional domains of the outer membrane proteins of Xac are analogous with Xcv’s.3. XacOmp4and XacOmp5of Xac both have signal peptide and OmpW domain, molecular weight of them are about20KDa.The length of amino acids and isoelectric point of XacOmp4and XacOmp5are204aa/5.90and207aa/6.28, respectively. The gene encoding XacOmp4and XacOmp5are612bp and638bp, respectively; XacOmp7has446aa, molecular weight of it is47KDa, isoelectric point of it is5.18and has signal peptide and TolueneX module. The gene encoding XacOmp7is1338bp; XacOmp12has365aa, molecular weight of it is39KDa, isoelectric point of it is4.63and has signal peptide and OmpA module. The gene encoding XacOmp12is1095bp.The four outer membrane proteins have high homology with others of Xanthomonas sp. We have cloned these four genes and expressed recombinant protein by IPTG induced.4. The functional testing of XacOmp7was used toluene as substrate and analyzing the toluene residual quantity by HPLC, confirming that XacOmp7accelerates the toluene catabolism.