Cloning, expression and characterization of L-arabinose isomerise from thermophilic Anoxybacillus kestanbolensis AC26Sari strain: Bioconversation of L-arabinose to L-ribulose

Abstract

L-Arabinose isomerase (L-AI) is a pivotal enzyme in the microbial pentose phosphate pathway. It is considered as a significant biological catalyst in rare sugar production. This enzyme can isomerize L-arabinose into L-ribulose and also D-galactose into D-tagatose. Here, we cloned the araA gene encoding L-arabinose isomerase from Anoxybacillus kestanbolensis AC26Sari strain, sequenced and over-expressed in E. coli BL21 (DE3): pLysS. This gene is involved in L-arabinose operon in A. kestanbolensis AC26Sari. DNA sequence analysis revealed an open reading frame of 1,506 bp, capable of encoding a polypeptide of 502 amino acid residues with calculated molecular weight of 55.6776 kDa. The recombinant was purified by heat treatment and Ni-HisTaq chromatography. The purified enzyme showed maximal activity at pH 8.5 and 65ºC and required divalent cations such as Co2+ and Mn2+ for its activity and thermostability. The apparent Km value of the enzyme for L-arabinose was 6.5 mM (Vmax, 140.1002 U/mg) as determined in the precence of both 1 mM Co2+ and Mn2+.