Development of Some Properties of a Thermophilic Recombinant Glucose Isomerase by Mutation

Dokuzparmak Ç., Colak A., Kolcuoglu Y., Akatin M., Ertunga N., Tuncay F.

APPLIED BIOCHEMISTRY AND MICROBIOLOGY, cilt.56, sa.2, ss.164-172, 2020 (SCI-Expanded) identifier identifier

  • Yayın Türü: Makale / Tam Makale
  • Cilt numarası: 56 Sayı: 2
  • Basım Tarihi: 2020
  • Doi Numarası: 10.1134/s0003683820020052
  • Dergi Adı: APPLIED BIOCHEMISTRY AND MICROBIOLOGY
  • Derginin Tarandığı İndeksler: Science Citation Index Expanded (SCI-EXPANDED), Scopus, Academic Search Premier, Agricultural & Environmental Science Database, BIOSIS, Biotechnology Research Abstracts, CAB Abstracts, Chemical Abstracts Core, EMBASE, Food Science & Technology Abstracts, Pollution Abstracts, Veterinary Science Database
  • Sayfa Sayıları: ss.164-172
  • Anahtar Kelimeler: glucose isomerase, thermophilic, Geobacillus, mutation, high fructose corn syrup, D-XYLOSE ISOMERASE, SINGLE-STEP PROCESSES, CLOSTRIDIUM-THERMOSULFUROGENES, PH OPTIMUM, PURIFICATION, CLONING, THERMOSTABILITY, EXPRESSION, STRAIN, SEQUENCE
  • Karadeniz Teknik Üniversitesi Adresli: Evet

Özet

Site-directed mutagenesis was made to obtain a glucose isomerase (GI) working at high temperatures and low pH values and having high affinity to the substrate and more pH and termal stability in comparison with the native enzyme. For this purpose, GI gene from Geobacillus caldoxylosilyticus TK4 was cloned to pET-28a(+) vector and H99Q, V184T and D102N mutations were performed. Biochemical features of the recombinant and mutant enzymes were identified. These mutations enabled the mutant enzymes to increase the optimum temperature and K-M and decrease the optimum pH and V-max values of the reaction. Furthermore, the mutant proteins had more thermal and pH stability than that of the recombinant GI. The mutant enzymes showed the highest activities in the presence of Co2+, Cu2+ and Mn2+ and were more resistant to some metals than the recombinant GI. In conclusion, the site-directed mutations led to improve the performance of the recombinant enzyme, which means that the mutant enzyme (TK4GI mutation) may be practicable in high fructose corn syrup production process.