| Mass expression of Apx I and Apx II of Actinobacillus pleuropneumoniae in Escherichia coli |
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Tae-Jung Kim, Bong-Joo Lee, Jae-Il Lee |
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College of Veterinary Medicine, Chonnam National University |
| 대장균에서 흉막폐렴균 독소 Apx I과 Apx II의 대량발현 |
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김태중, 이봉주, 이재일 |
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전남대학교 수의과대학 |
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| Abstract |
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Actinobacillus pleuropneumoniae is the causative agent of a porcine contagious pleuropneumonia. Among several virulence factors including exotoxin (Apx toxins), LPS, transferrin-binding proteins, OMPs, and some proteases, Apx toxins have been major targets for the protection study. In this study, cloning and expression of A. pleuropneumoniae Apx I and Apx II toxin, which are produced by all highly virulent strains, were performed by Escherichia coli expression system. Genes coding Apx I and II toxin were amplified from the A. pleuropneumoniae serotype 5 genomic DNA using polymerase chain reaction and cloned to a prokaryotic expression vector, pRSET. Expression of the Apx I and Apx II coding sequences in E. coli resulted in the formation of insoluble inclusion bodies purified according to a denaturing purification protocol, which employs the use of guanidium. Recombinant proteins were purified using $Ni^{2+}$-charged resin affinity purification. This expression and purification system made it possible to produce Apx I and Apx II in large amounts for further immunologic studies. |
| Key Words:
Actinobacillus pleuropneumoniae, Apx toxin, recombinant protein expression |
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