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Boosting heterologous protein production in transgenic dicotyledonous seeds using Phaseolus vulgaris regulatory sequences

Nature Biotechnology volume 20, pages 1265–1268 (2002)Cite this article

Abstract

Over the past decade, several high value proteins have been produced in different transgenic plant tissues such as leaves, tubers, and seeds1,2. Despite recent advances, many heterologous proteins accumulate to low concentrations, and the optimization of expression cassettes to make in planta production and purification economically feasible remains critical. Here, the regulatory sequences of the seed storage protein gene arcelin 5-I (arc5-I) of common bean (Phaseolus vulgaris)3 were evaluated for producing heterologous proteins in dicotyledonous seeds. The murine single chain variable fragment (scFv) G4 (ref. 4) was chosen as model protein because of the current industrial interest in producing antibodies and derived fragments in crops5,6. In transgenic Arabidopsis thaliana seed stocks, the scFv under control of the 35S promoter of the cauliflower mosaic virus (CaMV) accumulated to approximately 1% of total soluble protein (TSP). However, a set of seed storage promoter constructs boosted the scFv accumulation to exceptionally high concentrations, reaching no less than 36.5% of TSP in homozygous seeds. Even at these high concentrations, the scFv proteins had antigen-binding activity and affinity similar to those produced in Escherichia coli. The feasibility of heterologous protein production under control of arc5-I regulatory sequences was also demonstrated in Phaseolus acutifolius, a promising crop for large scale production.

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Figure 1: Outline of the T DNA constructs pParc5I-G4, pParc5I-Ω-G4, Pphas-G4, and P35S-G4 (not to scale).
Figure 2: Quantification and characterization of scFv extracted from seeds.

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Acknowledgements

We thank T. Hall (Texas A&M University) for providing the β-phaseolin promoter; Dominique Eeckhout, Peter Casteels, Bart Hoorelbeke, Ingrid Peck, Annelies De Clercq, Romain Uytterhaegen, Wilson Ardiles, Rebecca Verbanck, Lien Van Speybroeck, and Martine De Cock for advice and assistance; and the Vlaams Instituut voor de Bevordering van het Wetenschappelijk-Technologisch Onderzoek in de Industrie for financial support.

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Authors and Affiliations

  1. Department of Plant Systems Biology, Flanders Interuniversity Institute for Biotechnology, Ghent University, K.L. Ledeganckstraat 35, Gent, B 9000, Belgium

    Geert De Jaeger, Stanley Scheffer, Anni Jacobs, Mukund Zambre, Oliver Zobell, Alain Goossens & Ann Depicker

  2. Department of Applied Biological Sciences, Laboratory of Plant Genetics, Vrije Universiteit Brussel, Sint Genesius Rode, B 1640, Belgium

    Geert Angenon

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  1. Geert De Jaeger
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Correspondence to Ann Depicker.

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Jaeger, G., Scheffer, S., Jacobs, A. et al. Boosting heterologous protein production in transgenic dicotyledonous seeds using Phaseolus vulgaris regulatory sequences. Nat Biotechnol 20, 1265–1268 (2002). https://doi.org/10.1038/nbt755

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