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Expression of Acetohydroxyacid Synthase from Bacillus anthracis and Its Potent Inhibitors

  • Choi, Kyoung-Jae ;
  • Pham, Chien Ngoc ;
  • Jung, Hoe-Il ;
  • Han, Sung-Hwan ;
  • Choi, Jung-Do ;
  • Kim, Jin-Heung ;
  • Yoon, Moon-Young
  • Published : 2007.07.20

Abstract

Acetohydroxyacid synthase (AHAS, EC 2. 2. 1. 6) is the enzyme that catalyses the first step in the common pathway of the biosynthesis of the branched chain amino acids, valine, leucine and isoleucine. For the first time, the AHAS gene from Bacillus anthracis was cloned into the expression vector pET28a(+), and was expressed in the E. coli strain BL21(DE3). The purified enzyme was checked on 12% SDS-PAGE to be a single band with molecular weight of 65 kDa. The optimum pH and temperature for B. anthracis AHAS was at pH 7.5 and 37 oC, respectively. Kinetic parameters of B. anthracis were as follows: Km for pyruvate, K0.5 for ThDP and Mg2+ was 4.8, 0.28 and 1.16 mM respectively. AHAS from B. anthracis showed strong resistance to three classes of herbicides, Londax (a sulfonylurea), Cadre (an imidazolinone), and TP (a triazolopyrimidine). These results indicated that these herbicides could be used in the search for new anti-bacterial drugs.

Keywords

Acetohydroxyacid synthase;Herbicides;Resistance;Kinetic parameters

References

  1. Choi, K. J.; Noh, K. M.; Choi, J. D.; Park, J.; Won, H. S.; Kim, J. R.; Yoon, M. Y. Bull. Kor. Chem. Soc. 2006, 27, 1697-1700 https://doi.org/10.5012/bkcs.2006.27.10.1697
  2. Chang, A. K.; Duggleby, R. G. Biochem. J. 1997, 327, 161-169 https://doi.org/10.1042/bj3270161
  3. Spencer, R. C. Bacillus Anthracis, Int. Clin. Pathol. 2003, 56, 182-187 https://doi.org/10.1136/jcp.56.3.182
  4. Jerigan, J. A.; Stephens, D. S.; Ashford, D. A. Emerg. Infect. Dis. 2001, 7, 933-944 https://doi.org/10.3201/eid0706.010604
  5. Choi, K. J.; Yu, Y. G.; Hahn, H. G.; Choi, J. D.; Yoon, M. Y. FEBS Letter. 2005, 579, 4903-4910 https://doi.org/10.1016/j.febslet.2005.07.055
  6. Kil, M.; Chang, S. I. J. Biochem. Mol. Biol. 1998, 31, 287-293
  7. Poulsen, C.; Stougaard, P. Eur. J. Biochem. 1989, 185, 433-439 https://doi.org/10.1111/j.1432-1033.1989.tb15133.x
  8. Chang, A. K.; Duggleby, R. G. Biochem. J. 1997, 327, 161-169 https://doi.org/10.1042/bj3270161
  9. DeFelice, M.; Squires, C.; Levinthal, A. Biochem. Biophys. Acta 1978, 241, 9-17
  10. Karim, M.; Shim, M. Y.; Kim, J.; Choi, K. J.; Kim, J.; Choi, J. D.; Yoon, M. Y. Bull. Kor. Chem. Soc. 2006, 27, 549-555 https://doi.org/10.5012/bkcs.2006.27.4.549
  11. Khaleeli, N.; Li, R.; Townsend, C. A. J. Am. Chem. Soc. 1999, 121, 9223-9224 https://doi.org/10.1021/ja9923134
  12. Schloss, J. V.; Ciskanik, L. M.; Van Dyk, D. E. Nature 1988, 331, 360-362 https://doi.org/10.1038/331360a0
  13. Schloss, J. V.; Aulabaugh, A.; Barak, Z.; Chipman, D. M.; Schloss, J. V., Eds.; Biosynthesis of Branched Chain Amino Acids VCH press: Weinheim, Germany, 1988; pp 329-359
  14. Umbarger, H. E. In Synthesis of Amino Acid and Protein Arbstein, H. R. V., Ed.; MTP International Review of Science: Butterworths, London, 1975; pp 1-56
  15. Duggleby, R. G.; Pang, S. S. J. Biochem. Mol. Biol. 2000, 33, 1-36
  16. Davies, M. E. Plant Physiol. 1963, 39, 53-59 https://doi.org/10.1104/pp.39.1.53
  17. Singh, B. K.; Stidham, M. A.; Shaner, D. L. Anal. Biochem. 1988, 171, 173-179 https://doi.org/10.1016/0003-2697(88)90139-X

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