purification, and characterization of a thermophilic ribulokinase from Anoxybacillus kestanbolensis AC26Sari

Tokgoz M., Inan K., BELDÜZ A. O., GEDİKLİ Ö., ÇANAKÇI S.

TURKISH JOURNAL OF BIOLOGY, vol.38, no.5, pp.633-639, 2014 (SCI-Expanded) identifier identifier

  • Publication Type: Article / Article
  • Volume: 38 Issue: 5
  • Publication Date: 2014
  • Doi Number: 10.3906/biy-1401-92
  • Journal Indexes: Science Citation Index Expanded (SCI-EXPANDED), Scopus, TR DİZİN (ULAKBİM)
  • Page Numbers: pp.633-639
  • Karadeniz Technical University Affiliated: Yes


The gene encoding ribulokinase araB from Anoxybacillus kestanbolensis AC26Sari was cloned and sequenced. The recombinant protein was expressed in Escherichia coli BL21 under the control of isopropyl-beta-D-thiogalactopyranoside-inducible T7 promoter. The enzyme, designated as AC26RK, was purified with the MagneHis Protein Purification System. The molecular mass of the native protein, as determined by SDS-PAGE, was about 61 kDa. AC26RK was active throughout a broad pH (pH 5.0-10.0) and temperature (50-75 degrees C) range, and it had an optimum pH of 9.0 and optimum temperature of 60 C. The enzyme displayed about 90%-100% of its original activities after a 30-min incubation at a pH interval of 5.0-10.0. The enzyme exhibited a high level of D-ribulose activity with apparent K-m, V-max, and K-cat values of 0.94 mM, 3.197 U/mg, and 3.31 s(-1), respectively. AC26RK activity was strongly inhibited by Zn2+ but increased by Mg2+. The effects of some chemicals on the ribulokinase activity revealed that Anoxybacillus kestanbolensis AC26Sari does not need metallic cations for its activity. In this paper, we describe for the first time the cloning and characterization of a thermophilic ribulokinase from thermophilic bacteria.