Journal of Microbiology and Biotechnology

The Korean Society for Microbiology and Biotechnology (한국미생물·생명공학회)
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Constitutive Overexpression of the Endoxylanase Gene in Bacillus subtilis

  • Kim, Jong-Hyun (Department of Microbiology, Dong-Eui University) ;
  • Kim, Jung-Hoe (Department of Biologicla Sciences, Korea Advanced Institute of Science and Technology) ;
  • Kim, Sun-Chang (Department of Biologicla Sciences, Korea Advanced Institute of Science and Technology) ;
  • Nam, Soo-Wan (Department of Microbiology, Dong-Eui University)
  • Published : 2000.08.01
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Abstract

A strong constitutive $P_{JH}$ promoter from Bacillus was applied to overexpress the endoxylanase gene in B. subtilis. The expression plasmid, pHJKJ4, was designed to contain the $P_{JH}$ promoter and endoxylanase promoter ($P_B$), and introduced into B. subtilis DB104. Through batch fermentation of the trasformant cell on a maltose medium, endoxylanase was produced in a growth-associated manner as the predominant protein. The total activity reached about 600 unit/ml at the end of the cultivation, which corresponded to 698 mg endoxylanase protein/l with a specific activity of 860 unit/mg protein. It was also found that the segregational plasmid instability was less than 30% and most of the endoxylanase activity was detected in the culture medium. This result suggests that the secretory production of endoxylanase can be significantly enhanced with the use of the $P_{JH}$ promoter and high-cell density culture techniques, quantitatively as well as qualitatively.

Keywords

References

  1. Trends Biotechnol v.3 Microbiol xylanolytic systems Biely, P.
  2. Anal. Biochem v.72 A rpid and sensitive method for the quantification of microgrm quantites of protein utilizing the principle of protein-dye binding Bradford, M. M.
  3. Mol. Gen. Genet v.167 Characterization of plasmid transformation in Bacillus subtilis: Kinetic properties and the effect of DNA conformation Contente, S.;D. Dubnau
  4. Appl. Microbiol. Biotechnol v.50 High-elvel expression of an endoxylanase gene from Bacillus sp. in Bacillus subtilis DB104 for the production of xylobiose from xylan Jeong, K. J.;I. Y. Park;M. S. Kim;S. C. Kim
  5. Escherichia coli. Enzyme Microb. Technol v.22 Molecular cloning and characterization of an endoxylanase gene of Bacillus sp. Jeong, K. J.;P. C. Lee;I. Y. Park;M. S. Kim;S. C. Kim
  6. J. Bacteriol v.160 Construction of a Bacillus subtilis double mutant deficient in extracellular alkaline and neutral proteasa Kawamura, F.;R. H. Doi
  7. J. Microbiol. Biotechnol v.9 Expression of Bacillus macerans cyclodextrin glucanotrasferase in Bacillus subtilis Kim, C. S.;N. S. Han;D. H. Kweon;J. H. Seo
  8. Biotechnol. Lett v.15 Overproduction of extracellular endoglucanase by genetically engineered Bacillus subtilis Kim, J. H.;M. Y. Park
  9. J. Microbiol. Biotechnol v.8 Overproduction and secretion of B-glucosidase in Bacillus Kim, J. H.;B. R. Lee;M. Y. Park
  10. Nature v.227 Cleavage of structural proteins during the assembly of the head of bacteriophage T4 laemmli, U. K.
  11. Anal. Chem v.31 Use of dinitrosalicylic acid reagent for determination of reducing sugar Miller, G. L.
  12. J. Biol. Chem v.240 The release of enzymes from E. coli by osmotic shock and during the formation of spheroplast Neu, H. C.;L. A. Heppel
  13. FEMS Microbiol. Rev v.104 Molecular biology of xylan degradation Thomsom, J. A.
  14. Microbiol. Rev v.52 Multiplicity of B-1,4-xylanase in microorganism: Functions and applications Wong, K. K. Y.;U. L. lary;J. N. Saddler
  15. Biotechnol. Bioeng v.62 High-level secretory production of intact, biologically active staphylokinase from Bacillus subtilis Ye, R.;J. H. Kim;B. G. Kim;S. Szarka;E. Sihota;S. L. Wong