Overexpression of Nicotiana tabacum Acetolactate Synthase as an Inducible Fusion Protein in Escherichia coli: Production of a Polyclonal Antibody to Nicotiana tabacum Acetolactate Synthase

  • Chang, Soo-Ik (Department of Biochemistry, Chungbuk National University) ;
  • Kang, Moon-Kyeong (Department of Biochemistry, Chungbuk National University) ;
  • Kim, Hyun-Ju (Department of Biochemistry, Chungbuk National University) ;
  • Choi, Jung-Do (Department of Biochemistry, Chungbuk National University) ;
  • Namgoong, Sung-Keon (Department of Chemistry, Seoul Woman's University)
  • Received : 1996.05.28
  • Published : 1996.09.30


Acetolactate synthase (ALS, EC is the first common enzyme in the biosynthesis of leucine, isoleucine, and valine. It is the target enzyme for several classes of herbicides, including the sulfonylureas, the imidazolinones, the mazolopyrimidines, the pyrimidyl-oxy-benzoates, the pyrimidyl-thio-benzens, and the 4,6-dimethoxypyrimidines. An amino-terminal fragment of the sulfonylurea-resistant ALS gene (SurB) from Nicotiana tabaccum was cloned into the bacterial expression vector pGEX-2T. The resulting recombinant plasmid pGEX-ALS1 was used to transform Escherichia coli strain BL21, and the tobacco ALS was expressed in the bacteria as a protein fused with glutathione S-transferase (GST). Polyclonal antibodies against the fusion product (GST-ALS) were produced, and the sensitivity of GST-ALS with the rabbit anti-GST-ALS IgG was up to 50 ng. This antibody was used for Western blot analysis of the partially purified ALS from barley shoots. The results suggest that the polyclonal antibody produced in this study can be used to detect plant ALS.


acetolactate synthase;antibody;fusion protein;glutathione S-transferase;tobacco


  1. Proc. Natl. Acad. Sci. USA v.86 Smith, J.K.;Schloss, J.V.;Mazur, B.J. https://doi.org/10.1073/pnas.86.11.4179
  2. Gene v.69 Smith, D.B.;Johnson, K.S.
  3. Mol. Gen. Genet. v.219 Wiersma, P.A.;Schmiemann, M.G.;Condie, J.A.;Crosby, W.L.;Moloney, M.M. https://doi.org/10.1007/BF00259614
  4. Mol. Gen. Genet. v.224 Wiersma, P.A.;Hachey, J.E.;Crosby, W.L.;Moloney, M.M.
  5. Pest. Sci. v.31 Babczinski, P.;Zelinski, T. https://doi.org/10.1002/ps.2780310306
  6. Physiol. Plant. v.88 Bekkaoui, F.;Schorr, P.;Crosby, W. https://doi.org/10.1111/j.1399-3054.1993.tb01362.x
  7. J. Biol. Chem. v.29 Bernasconi, P.;Woodworth, A.R.;Rosen, B.A.;Subramanian, M.V.;Siehl, D.L.
  8. Science v.224 Chaleff, R.S.;Mauvaris, C.J. https://doi.org/10.1126/science.224.4656.1443
  9. Korean Biochem. J. (presently J. Biochem. Mol. Biol.) v.26 Choi, J.D.;Moon, H.;Chang, S.I.;Chae, J.K.;Shin, J.H.
  10. Z. Natturforsch. v.43c Dumer, J.;Boger, P.
  11. Pest. Sci. v.29 Gerwick, B.C.;Subramanian, M.V.;Loney-Gallant, V.;Chandler, D.P. https://doi.org/10.1002/ps.2780290310
  12. J. Biochem. Mol. Biol. (formerly Korean Biochem. J.) v.28 Kim, H.J.;Chang, S.I.
  13. Nature v.227 Laemmli, U.K. https://doi.org/10.1038/227680a0
  14. J. Biol. Chem. v.259 LaRossa, R.A.;Schloss, J.V.
  15. EMBO J. v.7 Lee, K.Y.;Townsend, J.;Tepperman, J.;Black, M.;Chui, C.F.;Mazur, B.;Dunsmuir, P.;Bedbrook, J.
  16. J. Biol. Chem. v.193 Lowry, O.H.;Rosebrough, N.J.;Farr, A.L.;Randall, R.J.
  17. Plant Physiol. v.85 Mazur, B.J.;Chui, C.F.;Smith, J.K. https://doi.org/10.1104/pp.85.4.1110
  18. Eur. J. Biochem. v.185 Poulsen, C.;Stougaard, P. https://doi.org/10.1111/j.1432-1033.1989.tb15133.x
  19. Plant Physiol. v.75 Ray, T.B. https://doi.org/10.1104/pp.75.3.827
  20. Science v.239 Saiki, R.K.;Gelfand, D.H.;Stoffel, S.;Scharf, S.J.;Higuchi, R.;Horn, G.T.;Mullis, K.B.;Erlich, H.A. https://doi.org/10.1126/science.2448875
  21. Molecular Cloning, A Laboratory Manual Sambrook, J.;Fritsch, E.F.;Maniatis, T.
  22. Biochemistry v.24 Scholoss, J.V.;Dyk, D.E.;Vasta, J.F.;Kytny, R.M. https://doi.org/10.1021/bi00339a034
  23. Plant Mol. Biol. v.76 Shaner, D.L.;Anderson, P.C.;Stidham, M.A.
  24. J. Biochem. Mol. Biol. (formerly Korean Biochem. J.) v.28 Shim, H.O.;Kim, D.W.;Chang, S.I.;Choi, J.D.
  25. Korean Biochem. J. (presently J. Biochem. Mol. Biol.) v.27 Sin, K.A.;Kim, H.J.;Choi, J.D.;Chang, S.I.
  26. Plant Physiol. v.97 Singh, B.;Schmitt, G.;Lillis, M.;Hand, J.M.;Misra, R. https://doi.org/10.1104/pp.97.2.657
  27. Plant Physiol. v.99 Singh, B.;Szamosi, I.;Hand, J.M.;Misra, R. https://doi.org/10.1104/pp.99.3.812