Study On Three Outer Membrane Proteins Of Xanthomonas Axonopodis Pv.citri XacOmp9,XacOmp11, XacOmp17

Citrus Canker is an important quarantine disease which influences citrus production of the world, and has a significant impact on the citrus industry. The pathogenic bacteria is Xanthomonas axonopodis pv.citri,(Xac). Outer membrane proteins are important components of the bacterial outer membrane, which play an important role in its life activities such as nutritional intake, substance metabolism and transport, maintaining the normal form of the bacteria, material synthesis and virulence of the bacteria, etc. Through the analysis of the Xac genome and proteome, we found that outer membrane protein is an important protein component of Xac. Meanwhile, there were few researchs on outer membrane proteins of Xac. Therefore, we guess that outer membrane protein has an important role in metabolism, virulence and material synthesis of Xac. Three outer membrane protein, XacOmp9, XacOmp11and XacOmp17, were selected to be analyzed with bioinformatics methods. Besides, the three genes were cloned and expressed to produce corresponding polyclonal antibodies. Through Western blot, indirect immunofluorescence positioning (IFA positioning), co-immunoprecipitation and LTQ mass spectrometry analysis, some results were obtained.The main results are as follows.1. XacOmp9gene is360bp in length, contains131amino acids with molecular weight14.9kDa. Its isoelectric point is9.81. It has an OmlA function module and no signal peptide. Through Indirect immunofluorescence localization(IFA), we found that XacOmp9protein was located on the surface of Xac cell. The co-immunoprecipitation and LTQ analysis showed that the coprecipitation proteins included biotin/sulfur octanoyl binding domain, regulatory proteins of two-component system, Hsp70and peptidyl proline cis-trans isomerase. Therefore, we predicted that XacOmp9adjust its sensitivity to SDS and antibiotics through the coordinating role with regulatory proteins of two-component system.2. XacOmpl1gene is462bp in length, contains181amino acids with molecular weight19.5kDa. Its isoelectric point is7.61. It has one signal peptide and an OmpA module. Through IFA positioning, we found that XacOmpl1protein was located on the surface of Xac cell. The co-immunoprecipitation and LTQ analysis showed that XacOmp11was mainly associated with some enzymes,such as Carboxypeptidase, Elongation factor G, GTP-binding protein lepA, tRNA synthetase, Gamma-glutamyl phosphate reductase, ATP synthase subunit alpha, ATP phosphoribosyltransferase, ribosomal protein, Chemotaxis response regulator.And these enzymes were mainly involved in the autologous protein synthesis and host protein hydrolysis. XacOmp11mainly use these oxidoreductases, protein hydrolyzate synthase enzymes to participate in the interaction against the host. In addition, OmpA precursor aversion requires the SecA protein and corresponding energy metabolism. XacOmp11may complete its drug resistance with chemotactic response regulator.In addition, the co-precipitation of XacOmp11found the common protein elongation factor G with XacOmp9. It shows that they are relevant to protein synthesis. But whether there was certain relationship between XacOmp9and XacOmp11,we needs for further research.3.XacOmp17gene is1464bp in length, contains488amino acids with molecular weight51.8kDa. Its isoelectric point is5.5. It has one signal peptide and two OEP function modules. IFA analysis shows that XacOmp17is located on the surface of Xac. Immunoprecipitation and LTQ analysis found that biotin/sulfur octanoyl, acetyl coenzyme A thiolase, TonB dependent receptors may be associated with XacOmp17.And these further support the function of XacOmp17in the formation of trimeric pore and matrix transport.Hypothetical protein XAC0019assoiated with XacOmp17, relate to the degradation of toluene and aromatic compounds. We surmise XacOmp17provides access for the degradation of toluene and aromatic compounds to enter cells, in order to make hypothetical protein XAC0019further play the role of toluene degradation. However, whether the XacOmp17have multi-drug resistance, we need further research.