Expression of the C-terminal of 34kDa protein of Mycobacterium paratuberculosis

Mycobacterium paratuberculosis의 34kDa C-terminal 단백질의 발현

  • Kim, Doo (Department of Veterinary Medicine, Kangwon National University) ;
  • Park, Hyung-wook (Department of Veterinary Medicine, Kangwon National University)
  • 김두 (강원대학교 수의학과) ;
  • 박형욱 (강원대학교 수의학과)
  • Received : 2000.01.08
  • Published : 2000.03.25

Abstract

Paratuberculosis (Johne's disease), a chronic enteritis produced by Mycobacterium paratuberculosis, affects a large proportion of ruminants in all continents and causes important economic losses. The identification of well-characterized and species-specific components of M paratuberculosis would provide the means to improve the specificity and sensitivity of immunodiagnostic assays for Johne's disease. The aims of this study were to express the recombinant C-terminal of 34kDa protein (rC34P) of M paratuberculosis in E coli and to investigate the effectiveness of this protein in detecting antibodies to the native protein in sera from paratuberculosis infected cattle. The C-terminal of the gene encoding the 34kDa protein was amplified by polymerase chain reaction from the chromosomal DNA of M paratuberculosis (ATCC 19698) and cloned into vector pGEX-4T-2. Then, cloned plasmid was transformed into E coli DH5${\alpha}$ and the rC34P was overexpressed. The rC34P was purified by affinity chromatography and gel filtration. The rC34P was examined antigenicity by Western blot. The rC34P was reactive with culture positive bovine serum and hyperimmune rabbit anti-M paratuberculosis serum but was not reactive with culture negative bovine serum and tuberculin positive bovine serum in Western blot. In conclusion, the rC34P produced in this study is expected as a useful candidate for antigen in serological diagnosis of Johne's disease.