Activity and thermostability increase of xylanase following transplantation with modules sub-divided from hyper-thermophilic CBM9_l-2

摘要

Transplantation is useful for elucidating the functions of structural modules and for engineering enzyme properties. Unexpectedly, transplanting a hyper-thermophilic carbohydrate-binding module, CBM9.1 -2, into the mesophilic Aspergillus niger GH11 xylanase (Xyn) slightly decreased the thermal inactivation half-life of Xyn. This effect was further investigated by dividing the CBM9-1 -2 module into two smaller parts, Cl and C2, which were transplanted into Xyn to create the chimeras Xyn-C1 and Xyn-C2. Both chimeras exhibited higher catalytic activities on xylan than native Xyn. Xyn-C2 exhibited higher binding affinities for both oat spelt and birch wood xylans, and its thermal inactivation half-life (69.3 min) was 4 or 5 times longer than that of Xyn (17.6 min), Xyn-Cl (13.4min), and the original chimera containing CBM9_1-2 (13.8min). In contrast, Xyn-Cl exhibited higher binding affinity for oat spelt xylan, but not for birch wood xylan. Through this rational engineering of the fungal xylanase, the C2 sub-module was shown to have a different thermostabilizing effect than the Cl sub-module. The different functions of the smaller parts of a large module can play pivotal roles in transplantation.