the expression of a xylanase targeted to er-protein bodies provides a simple strategy to produce active insoluble enzyme polymers in tobacco plants木聚糖酶的表达针对er-protein机构提供了一个简单的战略产生积极的不溶性酶聚合物在烟草植物.pdf

The Expression of a Xylanase Targeted to ER-Protein Bodies Provides a Simple Strategy to Produce Active Insoluble Enzyme Polymers in Tobacco Plants 1,2 1 1 1 Immaculada Llop-Tous , Miriam Ortiz , Margarita Torrent , M. Dolors Ludevid * 1 Department of Molecular Genetics, Centre for Research in Agricultural Genomics (CRAG, Consortium CSIC-IRTA-UAB), Barcelona, Spain, 2 ERA Biotech, Barcelona, Spain Abstract Background: Xylanases deserve particular attention due to their potential application in the feed, pulp bleaching and paper industries. We have developed here an efficient system for the production of an active xylanase in tobacco plants fused to a proline-rich domain (Zera) of the maize storage protein c-zein. Zera is a self-assembling domain able to form protein aggregates in vivo packed in newly formed endoplasmic reticulum-derived organelles known as protein bodies (PBs). Methodology/Principal Findings: Tobacco leaves were transiently transformed with a binary vector containing the Zera- xylanase coding region, which was optimized for plant expression, under the control of the 35S CaMV promoter. The fusion protein was efficiently expressed and stored in dense PBs, resulting in yields of up to 9% of total protein. Zera-xylanase was post-translationally modified with high-mannose-type glycans. Xylanase fused to Zera was biologically active not only when solubilized from PBs but also in its insoluble form. The resistance of insoluble Zera-xylanase to trypsin digestion demonstrated that the correct folding of xylanase in PBs was not impaired by Zera oligomerization. The activity of insoluble Zera-xylanase was enhanced when substrate accessibility was facilitated by