Tomato cystatin SlCYS8 as a stabilizing fusion partner for human serpin expression in plants

Sainsbury, Frank, Varennes-Jutras, Philippe, Goulet, Marie-Claire, D'Aoust, Marc-Andr and Michaud, Dominique (2013) Tomato cystatin SlCYS8 as a stabilizing fusion partner for human serpin expression in plants. Plant Biotechnology Journal, 11 9: 1058-1068. doi:10.1111/pbi.12098

Author Sainsbury, Frank
Varennes-Jutras, Philippe
Goulet, Marie-Claire
D'Aoust, Marc-Andr
Michaud, Dominique
Title Tomato cystatin SlCYS8 as a stabilizing fusion partner for human serpin expression in plants
Journal name Plant Biotechnology Journal   Check publisher's open access policy
ISSN 1467-7644
Publication date 2013-01-01
Year available 2013
Sub-type Article (original research)
DOI 10.1111/pbi.12098
Open Access Status DOI
Volume 11
Issue 9
Start page 1058
End page 1068
Total pages 11
Place of publication Oxford, United Kingdom
Publisher Wiley-Blackwell
Language eng
Abstract Studies have reported the usefulness of fusion proteins to bolster recombinant protein yields in plants. Here, we assess the potential of tomato SlCYS8, a Cys protease inhibitor of the cystatin protein superfamily, as a stabilizing fusion partner for human alpha-1-antichymotrypsin (α1ACT) targeted to the plant cell secretory pathway. Using the model expression platform Nicotiana benthamiana, we show that the cystatin imparts a strong stabilizing effect when expressed as a translational fusion with α1ACT, allowing impressive accumulation yields of over 2 mg/g of fresh weight tissue for the human serpin, a 25-fold improvement on the yield of α1ACT expressed alone. Natural and synthetic peptide linkers inserted between SlCYS8 and α1ACT have differential effects on protease inhibitory potency of the two protein partners in vitro. They also have a differential impact on the yield of α1ACT, dependent on the extent to which the hybrid protein may remain intact in the plant cell environment. The stabilizing effect of SlCYS8 does not involve Cys protease inhibition and can be partly reproduced in the cytosol, where peptide linkers are less susceptible to degradation. The effect of SlCYS8 on α1ACT yields could be explained by: (i) an improved translation of the human protein coding sequence; and/or (ii) an overall stabilization of its tertiary structure preventing proteolytic degradation and/or polymerization. These findings suggest the potential of plant cystatins as stabilizing fusion partners for recombinant proteins in plant systems. They also underline the need for an empirical assessment of peptide linker functions in plant cell environments.
Keyword Alpha-1-antichymotrypsin
Fusion proteins
Peptide linkers
Plant cystatins
Plant-based protein expression
Recombinant proteins
Q-Index Code C1
Q-Index Status Confirmed Code
Institutional Status UQ

Document type: Journal Article
Sub-type: Article (original research)
Collections: Official 2014 Collection
Australian Institute for Bioengineering and Nanotechnology Publications
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Citation counts: TR Web of Science Citation Count  Cited 11 times in Thomson Reuters Web of Science Article | Citations
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