Expression and characterization of an endoxylanase in Escherichia coli and Bacillus subtilis

Abstract

The gene encoding a pre-endoxylanase of Bacillus sp. has been cloned and expressed in Escherichia coli and Bacillus subtilis. The pre-endoxylanase consisted of 28 amino acid-signal peptide and 213 amino acid endoxylanase. The signal peptide of 28-amino acid was processed both in E. coli and B. subtilis. The endoxylanase expressed in B. subtilis was identical to that expressed in E. coli. To increase the expression level of the endoxylanase in B. subtilis, a dimeric endoxylanase gene was constructed in a tandem unit using a gene amplification system. However, the dimeric endoxylanase gene cloned in rolling-cicle replicating plasmid spontaneously rearranged to a monomeric one in B. subtilis. To solve the plasmid instability in B. subtilis, the theta (Θ)-replicating plasmid was substituted for rolling-circle replicating one. The dimeric endoxylanase gene cloned in the theta (Θ)-replicating plasmid remained structurally stable in B. subtilis. Comparative analysis of endoxylanase expression revealed that the production of the endoxylanase in B. subtilis containing a dimeric endoxylanase gene was 50% greater than that of B. subtilis containing a monomeric one.