Protection of Recombinant Mammalian Antibodies from Development-Dependent Proteolysis in Leaves of Nicotiana benthamiana

Robert S., Khalf M., Goulet M.-C., D'Aoust M.-A., Sainsbury F. and Michaud D. (2013) Protection of Recombinant Mammalian Antibodies from Development-Dependent Proteolysis in Leaves of Nicotiana benthamiana. PLoS ONE, 8 7: e70203.1-e70203.9. doi:10.1371/journal.pone.0070203


Author Robert S.
Khalf M.
Goulet M.-C.
D'Aoust M.-A.
Sainsbury F.
Michaud D.
Title Protection of Recombinant Mammalian Antibodies from Development-Dependent Proteolysis in Leaves of Nicotiana benthamiana
Journal name PLoS ONE   Check publisher's open access policy
ISSN 1932-6203
Publication date 2013-07
Year available 2013
Sub-type Article (original research)
DOI 10.1371/journal.pone.0070203
Open Access Status DOI
Volume 8
Issue 7
Start page e70203.1
End page e70203.9
Total pages 9
Place of publication San Francisco, United States
Publisher Public Library of Science (PLoS)
Collection year 2014
Language eng
Formatted abstract
The expression of clinically useful proteins in plants has been bolstered by the development of high-yielding systems for transient protein expression using agroinfiltration. There is a need now to know more about how host plant development and metabolism influence the quantity and quality of recombinant proteins. Endogenous proteolysis is a key determinant of the stability and yield of recombinant proteins in plants. Here we characterised cysteine (C1A) and aspartate (A1) protease profiles in leaves of the widely used expression host Nicotiana benthamiana, in relation with the production of a murine IgG, C5-1, targeted to the cell secretory pathway. Agroinfiltration significantly altered the distribution of C1A and A1 proteases along the leaf age gradient, with a correlation between leaf age and the level of proteolysis in whole-cell and apoplast protein extracts. The co-expression of tomato cystatin SlCYS8, an inhibitor of C1A proteases, alongside C5-1 increased antibody yield by nearly 40% after the usual 6-days incubation period, up to ∼3 mg per plant. No positive effect of SlCYS8 was observed in oldest leaves, in line with an increased level of C1A protease activity and a very low expression rate of the inhibitor. By contrast, C5-1 yield was greater by an additional 40% following 8- to 10-days incubations in younger leaves, where high SlCYS8 expression was maintained. These findings confirm that the co-expression of recombinant protease inhibitors is a promising strategy for increasing recombinant protein yields in plants, but that further opportunity exists to improve this approach by addressing the influence of leaf age and proteases of other classes.
Q-Index Code C1
Q-Index Status Provisional Code
Institutional Status Non-UQ

Document type: Journal Article
Sub-type: Article (original research)
Collections: Non HERDC
Australian Institute for Bioengineering and Nanotechnology Publications
 
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