Biochemical characterization of a novel thermostable feruloyl esterase from Geobacillus thermoglucosidasius DSM 2542(T)

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Sal F., Colak D. N., Güler H. İ., Çanakçı S., Beldüz A. O.

MOLECULAR BIOLOGY REPORTS, cilt.46, sa.4, ss.4385-4395, 2019 (SCI-Expanded) identifier identifier identifier

  • Yayın Türü: Makale / Tam Makale
  • Cilt numarası: 46 Sayı: 4
  • Basım Tarihi: 2019
  • Doi Numarası: 10.1007/s11033-019-04893-6
  • Dergi Adı: MOLECULAR BIOLOGY REPORTS
  • Derginin Tarandığı İndeksler: Science Citation Index Expanded (SCI-EXPANDED), Scopus
  • Sayfa Sayıları: ss.4385-4395
  • Anahtar Kelimeler: Geobacillus, Feruloyl esterase, Recombinant, Thermostable, ASPERGILLUS-NIGER, ACID, PURIFICATION, IDENTIFICATION, SPECIFICITY, CLONING, ENZYME
  • Karadeniz Teknik Üniversitesi Adresli: Evet

Özet

The ferulic acid esterase (FAE) gene from Geobacillus thermoglucosidasius DSM 2542(T) was cloned into pET28a(+) expression vector and characterized and is being reported in this study for the first time in Geobacillus. The enzyme, designated as GthFAE, was purified by heat shock and ion-exchange column chromatography. In addition, a second clone containing a Histidine tag was expressed and purified by affinity column chromatography demonstrating future potential for scale-up. FAE gene contains an open reading frame (ORF) of 759-bp encoding a hypothetical 252 amino acid protein, a molecular mass of 28.11kDa and an isoelectric point of 5.53. From this study it was found that GthFAE had optimal activity at 50 degrees C and pH of 8.5. Furthermore, the enzyme has been found to retain 64% of its activity after two days incubation at 50 degrees C and exhibited a high level of functionality with p-nitrophenyl caprylate (C8). K-m, V-max, k(cat) and k(cat)/K-m values for p-nitrophenyl caprylate were determined as 0.035mM, 11,735 mu mol/min/mg protein, 5491 (1/s) and 156,885s(-1)mM(-1) respectively. The combination of higher activity and stability compared to previously reported FAEs makes GthFAE a potential candidate for use in the paper manufacturing industry.