High-level soluble expression of a thermostable xylanase from thermophilic fungus Thermomyces lanuginosus in Escherichia coli via fusion with OsmY protein

• We constructed an expression vector pET-OsmY-xynA.
• The fusion protein was expressed as soluble form without IPTG induction.
• Induction of the fusion protein is due to auto-induction.
• The fusion protein was purified and characterized.

A thermostable xylanase is encoded by xynA from fungus Thermomyces lanuginosus. The problem emerged from overexpression of xynA in Escherichia coli has been the formation of inclusion bodies. Here we describe the xynA was fused with the hyperosmotically inducible periplasmic protein of E. coli, OsmY. The fusion protein OsmY-xynA was expressed as almost all soluble form. The soluble expression level of fusion protein reached 98 ± 6 U/ml when cells containing pET-OsmY-xynA were expressed without IPTG induction at 37 °C. The induction is probably due to auto-induction due to lactose in the medium (Studier (2005) [21]). The cells harboring pET-OsmY-xynA expressed an activity level about 24 times higher than that expressed from pET-20b-xynA. Xylanase activity was observed in the extracellular (36 ± 1.3 U/ml) and the periplasmic (42 ± 4 U/ml) when cells containing pET-OsmY-xynA were induced without IPTG addition. After the cold osmotic shock procedure followed by nickel affinity chromatography, the purified fusion protein showed a single band on SDS–PAGE gel with a molecular mass of 44 kDa. The purified fusion enzyme exhibited the highest activity at 65 °C and pH 6.0.