Prokaryotic Expression and Secretion Characterization of Annexin B5,B15,and B25 from Echinococcus granulosus
This study aims to investigate the immunoreactivity and secretion characteristics of the Echinococcus granulosus(E.granulosus)annexin B5,B15,and B25 in the intermediate host tis-sues,laying the foundation for further research on the interaction between annexin and hosts.In this study,total RNA was extracted from the protoscolex of E.granulosus,and then reverse transcribed into cDNA.The full-length coding sequences of EgANXB5,EgANXB15,and EgANXB25 genes were amplified by PCR using the cDNA as a template.The recombinant plas-mids pET32a-Eg ANXB5,pET32a-Eg ANXB15,and pET32a-Eg ANXB25 were constructed using homologous recombination method,and the recombinant proteins were expressed in Esche-richia coli.The immunoreactivity of these recombinant proteins was analyzed using Western blotting,and their secretion in the intermediate host tissues was examined by immunofluores-cence staining.The recombinant EgANXB5,EgANXB15,and EgANXB25 were successfully cloned and expressed.The coding sequence(CDS)of EgANXB25 was corrected(GenBank:OR245515.1).These three proteins could be specifically recognized by positive canine sera for E.granulosus and positive mouse sera for E.granulosus,indicating that they had strong immuno-reactivity.Moreover,immunofluorescence staining revealed that EgANXB5,EgANXB15,and EgANXB25 were primarily distributed in the liver parenchyma near the E.granulosus cyst.Re-combinant EgANXB5,EgANXB15,and EgANXB25 exhibite strong immunoreactivity and these proteins have the potential to secrete into the hepatic parenchymal tissue surrounding the cyst,possibly further participating in the interaction between E.granulosus and their host.
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