Characterization of a xyloglucananse in biodegradation of woody plant xyloglucan from Caldicellulosiruptor kronotskyensis

Abstract

The 2,835-bp open reading frame of ckxgl74A (Locus_tag CALKRO_RS04315) with a natural carbohydrate module (CBM3b) from thermophilic anaerobic microorganism Caldicellulosiruptor kronotskyensis encodes a calculated 104-kDa of GH74 xyloglucanase Ckxgl74A. The purified recombinant Ckxgl74A expressed in Escherichia coli BL21 (DE3) revealed its optimal pH of 4.5 and temperature of 80 °C. The Ckxgl74A was stable over a temperature no more than 70 °C and a pH range of 4.5 to 5.0. Kinetic experiments with xyloglucan as a substrate gave a Km of 2.29 ± 0.04 mg mL-1, Vmax of 22.98 ± 0.02 mol mg-1 min-1, and kcat of 66.98 ± 0.01 s-1. Its activity could be activated by Ca2+ approximately two folds, while being significantly inhibited by Cu2+. These results showed that Ckxgl74A could be utilized in acid condition and possessed a good thermostability.