Molecular Cloning and Expression of an Endo-xylanase Gene from Bacillus stearothermuphilus into Escherichia coli

Bacillus stearothermophilus로 부터 Endo-xylanase 유전자의 클로닝 및 Escherichia coli에서의 발현

  • 조상구 (고려대학교 자연과학대학 유전공학과) ;
  • 박성수 (고려대학교 자연과학대학 유전공학과) ;
  • 박영인 (고려대학교 자연과학대학 유전공학과) ;
  • 최용진 (고려대학교 자연과학대학 유전공학과)
  • Published : 1992.06.01

Abstract

Genomic DNA of Bacillus stearothemzophilus, which expressed alkalophilic and thermophilic xylanases, was partially digested with HindIII, cloned into pBR322, and subsequently transferred into the Escherichra coli HB101 cells. Three among 5, 000 transformants screened formed clear zones around their colonies. From the functional clones, three recombinant plasmids (pMG11, pMG12 and pMG13) had been isolated, and they were identified to carry the same 4 kb HindIII fragment originated from B. stearothemzophilus which was responsible for the xylanase activity. pMGl3, however, had the foreign DNA of opposite orientation compared to the other two recombinant plasmids. This recombinant plasmid gave much lower xylanase activity. B. stearothermophilus was observed to produce at least three xylanase activities as evidenced by the PAGE-xylan zymogram. The xylanase from E. coli HB101/pMG12 was judged to correspond to the largest among the three B. stearothermophilus xylanases observed in the zymogrom. The enzyme hydrolyzed xylooligosaccharides larger than xylotriose and degraded xylan to produce xylobiose and xylotriose as major products. The xylanase was considered to have trans-xylosidase activity, too.

Keywords

Cloning;endo-xylanase;B. stratothermophillus;zymogram;tran-xylosidase activity