Comparative study of the subcellular localisation and the N-glycosylation patterns of a recombinant glycoprotein expressed in Oryza sativa, Nicotina tabacum and Medicago truncatula

Nowadays, plants have become a promising alternative over the traditional expression systems for the production of biologically active complex recombinant proteins. So far, research has been focussed on increasing accumulation levels of recombinant proteins expressed in many plant models. On the other hand, the cell compartment where recombinant proteins are deposited often refers to protein quality in term of protein folding and post-translational modification (e.g. N-glycosylation). Recombinant proteins directed to the secretory pathway in plants require a leader peptide for translocation into the endoplasmic reticulum. In the absence of further targeting information such proteins are generally secreted, via the default pathway, to the apoplast. This has been well-documented in protoplasts and leaf cells, but the trafficking of recombinant proteins in storage tissues such as endosperm or cotyledons has only received little attention. In addition, it has been suggested that the high specificity of seed tissue for starch, minerals and proteins storage might interfere with protein secretion. We used Aspergillus niger phytase as a model glycoprotein to compare the intracellular fate and the N-glycosylation profile of a recombinant protein in different plant tissues (seeds and leaves) in rice, tobacco and Medicago truncatula. After determination of N-glycosylation patterns and microscopy analyses, we reported that the recombinant protein was preferably retained in protein storage organelles within rice and tobacco endosperm cells, while it was efficiently secreted in leaf and cotyledon tissues as expected for protein following the default secretory pathway. Interestingly, in rice and M. tuncatula leaves, a significant part of A. niger phytase was misrouted to the vacuole. Besides, we revealed an important heterogeneity in the N-glycosylation patterns of A. niger phytase when produced in different plants and tissues. In addition, only A. niger phytase extracted from M. truncatula leaves harboured Lewisa-bearing N-glycans known to be potentially highly immunogenic. Together, these results showed that localisation and N-glycosylation of a recombinant protein is tissue dependent, with obvious implications for the production of pharmaceutical proteins by molecular farming.

Grantor dc:publisher

Publikationsserver der RWTH Aachen University

Year dc:date

2005

Author dc:creator

• Marcel, Sylvain

Contributors dc:contributor

• Fischer, Rainer

• info:eu-repo/classification/ddc/580
• Pflanzen (Botanik)
• protein deposition
• N-glycosylation
• Aspergillus niger phytase

Statement dc:rights

• info:eu-repo/semantics/openAccess

Language dc:language

eng

Identifier

https://publications.rwth-aachen.de/search?p=id:%22RWTH-CONV-123729%22

OAI identifier oai:identifier

oai:publications.rwth-aachen.de:62136