Expression of a thermostable restriction endonuclease in recombinant Escherichia coli cells and optimization of fermentation conditions

Toksoy E., Onsan Z., Kirdar B.

WORLD JOURNAL OF MICROBIOLOGY & BIOTECHNOLOGY, cilt.18, sa.1, ss.23-27, 2002 (SCI-Expanded) identifier identifier

  • Yayın Türü: Makale / Tam Makale
  • Cilt numarası: 18 Sayı: 1
  • Basım Tarihi: 2002
  • Doi Numarası: 10.1023/a:1013921713072
  • Dergi Adı: WORLD JOURNAL OF MICROBIOLOGY & BIOTECHNOLOGY
  • Derginin Tarandığı İndeksler: Science Citation Index Expanded (SCI-EXPANDED), Scopus
  • Sayfa Sayıları: ss.23-27
  • Anahtar Kelimeler: Escherichia coli, expression, fermentation, optimization, restriction endonuclease, Taq I, HIGH-LEVEL EXPRESSION, TAQI RESTRICTION, SYSTEM
  • Marmara Üniversitesi Adresli: Evet

Özet

Taq I restriction endonuclease gene of the thermophilic eubacterium Thermus aquaticus YT-1 (ATCC 25104) was successfully cloned and expressed in recombinant Escherichia coli cells under the control of the lac promoter/operator system. Higher Taq I endonuclease specific activities and biomass yields were obtained from E. coli ER2508(pUCTaq) cells when they were induced at the late-exponential phase of their growth. Taq I endonuclease expression was found to be host strain-dependent such that, among the three different strains examined, E. coli XL1(pUCTaq) produced the highest specific Taq I endonuclease activities for longer induction periods. Decreasing the inducer concentration from 1 to 0.1 mM not only improved the specific enzyme activity yields but also is more economical, considering the high cost of isopropyl-beta-D-thiogalactopyranoside (IPTG). The optimum culture temperature was found to be 37 degreesC. Taq I endonuclease specific activity recovered from E. coli XL1(pUCTaq) cells was 935 U/mg under optimum conditions.