Irons, Sarah L., Nuttall, J. (James), Floss, Doreen M., Frigerio, Lorenzo, Kotzer, Amanda M. and Hawes, C. R.. (2008) Fluorescent protein fusions to a human immunodeficiency virus monoclonal antibody reveal its intracellular transport through the plant endomembrane system. Plant Biotechnology Journal, Vol.6 (No.7). pp. 649-662. ISSN 1467-7644

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Abstract

In order to further understand the production and intracellular trafficking of pharmaceutical proteins in plants, the light and heavy chains (LC and HC) of the human immunodeficiency virus neutralizing monoclonal antibody 2G12 were fused to fluorescent proteins [Venus and monomeric red fluorescent protein (mRFP)] to enable the visualization of their passage through the plant cell. Co-expression of LC and HC with various markers of the endomembrane system demonstrated that LC fusions were found in mobile punctate structures, which are likely to be pre-vacuolar compartments (PVCs) as a proportion of the LC fusions were found to be located in the vacuole. In addition, apoplast labelling was also observed with a 2G12LC-RFP fusion. The HC fusion expressed alone was found only in the endoplasmic reticulum (ER). When the LC and HC fusions were expressed together, they were found to co-locate to larger punctate structures, which were morphologically distinct from any observed on expression of LC or HC alone. These structures appeared to be in close association with the ER and their labelling partially overlapped with PVC marker fluorescence, but no increase in apoplast labelling was observed. Co-immunoprecipitation data demonstrated that the presence of the fluorescent proteins did not affect the assembly of the antibody, and also showed the association of BiP with the antibody chains. The antigen-binding activity of the Venus-fused 2G12 antibody was confirmed by enzyme-linked immunosorbent assay.

Item Type:	Journal Article
Subjects:	Q Science > QK Botany Q Science > QR Microbiology > QR180 Immunology
Divisions:	Faculty of Science > Life Sciences (2010- )
Library of Congress Subject Headings (LCSH):	Plant proteins -- Physiological transport, Fluorescence, Proteins, Monoclonal antibodies, Pharmaceutical biotechnology, HIV (Viruses)
Journal or Publication Title:	Plant Biotechnology Journal
Publisher:	Wiley-Blackwell Publishing Ltd.
ISSN:	1467-7644
Official Date:	September 2008
Volume:	Vol.6
Number:	No.7
Number of Pages:	14
Page Range:	pp. 649-662
Identification Number:	10.1111/j.1467-7652.2008.00348.x
Status:	Not Peer Reviewed
Publication Status:	Published
Access rights to Published version:	Restricted or Subscription Access
URI:	http://wrap.warwick.ac.uk/id/eprint/29532

Data sourced from Thomson Reuters' Web of Knowledge

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