Cloning, expression and characterization of a cold-adapted endo-1, 4--glucanase from A1, a symbiotic bacterium of .

Background: Many biotechnological and industrial applications can benefit from cold-adapted EglCs through increased efficiency of catalytic processes at low temperature. In our previous study, A1 which was isolated from a wood-inhabiting termite could secrete a cold-adapted EglC. However, its EglC was difficult to purify for enzymatic properties detection because of its low activity (0.8 U/ml). The objective of the present study was to clone and express the gene in to improve production level and determine the enzymatic properties of the recombinant enzyme.

Methods: The gene was cloned from A1 by thermal asymmetric interlaced PCR. was transformed into vector pET22b and functionally expressed in . The recombination protein EglC22b was purified for properties detection.

Results: SDS-PAGE revealed that the molecular mass of the recombinant endoglucanase was approximately 42 kDa. The activity of the pET22b-EglC crude extract was 9.5 U/ml. Additionally, it was active at pH 6.5-8.0 with an optimum pH of 7.0. The recombinant enzyme had an optimal temperature of 30-40 °C and exhibited >50% relative activity even at 5 °C, whereas it lost approximately 90% of its activity after incubation at 60 °C for 30 min. Its activity was enhanced by Co and Fe, but inhibited by Cd, Zn, Li, Triton X-100, DMSO, acetonitrile, Tween 80, SDS, and EDTA.

Conclusion: These biochemical properties indicate that the recombinant enzyme is a cold-adapted endoglucanase that can be used for various industrial applications.