Cloning and Expression of an Oligo-1,6-glucosidase Gene from Arthrobacter globiformis I42 and Biochemical Characterization of the Recombinant Enzyme

Kouzou Yamaguchi, Naoki Morimoto, Yi Wang, Kenji Watanabe, Takehiro Unno, Hiroyuki Ito, Hirokazu Matsui
2004 Journal of Applied Glycoscience  
The gene encoding an oligo-1,6-glucosidase was cloned in terms of walking downstream from the glucodextranase gene of the chromosomal DNA of Arthrobacter globiformis I42. An open reading frame consisted of 1731 base pairs that encoded a mature protein of 577 amino acids (Mr, 63,000) was found. Transformed Escherichia coli cells carrying the 1.7-kb fragment overproduced the oligo-1,6-glucosidase under control of the T7 promoter of a pET system. Kinetic analyses of the recombinant protein gave Km
more » ... ant protein gave Km 1.76 mM and k0 697 s 1 for p-nitrophenyl α-D-glucopyranoside and Km 24.1 mM and k0 41 s 1 for isomaltose. Its deduced amino acid sequence showed 54% similarity to two amino acid sequences of Bacillus cereus oligo-1,6glucosidase and Bacillus sp. α-glucosidase. The oligo-1,6-glucosidase has four conserved regions shared with αamylases. The gene cluster consisted of the glucodextranase and oligo-1,6-glucosidase genes, suggesting that both genes could participate in the degradation for utilization of dextran in the bacterium.
doi:10.5458/jag.51.37 fatcat:ai6xz6xx2jej5bljvaqypvxrpa