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High-yield production of a human therapeutic protein in tobacco chloroplasts

Nature Biotechnology volume 18, pages 333–338 (2000)Cite this article

Abstract

Transgenic plants have become attractive systems for production of human therapeutic proteins because of the reduced risk of mammalian viral contaminants, the ability to do large scale-up at low cost, and the low maintenance requirements. Here we report a feasibility study for production of a human therapeutic protein through transplastomic transformation technology, which has the additional advantage of increased biological containment by apparent elimination of the transmission of transgenes through pollen. We show that chloroplasts can express a secretory protein, human somatotropin, in a soluble, biologically active, disulfide-bonded form. High concentrations of recombinant protein accumulation are observed (>7% total soluble protein), more than 300-fold higher than a similar gene expressed using a nuclear transgenic approach. The plastid-expressed somatotropin is nearly devoid of complex post-translational modifications, effectively increasing the amount of usable recombinant protein. We also describe approaches to obtain a somatotropin with a non-methionine N terminus, similar to the native human protein. The results indicate that chloroplasts are a highly efficient vehicle for the potential production of pharmaceutical proteins in plants.

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Figure 1: Analysis of transplastomic plants carrying recombinant hST genes integrated into the plastid genome.
Figure 2: Accumulation of hST protein in transplastomic lines.
Figure 3: Recombinant chloroplast hST contains the correct disulfide bonds.
Figure 4: Plastid-expressed hST is biologically active.
Figure 5: Chloroplast transgenes are maternally inherited.

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Acknowledgements

We thank Robin Weinberg for a ubiquitin–hST fusion plasmid, Titik Dian for assistance with protein purification, Dawn Dufield for mass spectrometry analysis, Skooter Jennings for amino acid analysis, and Pal Maliga, Rutgers University, for the pPRV plasmids. We also thank Michael Montague for a critical reading of the manuscript and Ganesh Kishore, Michael Montague, Julio Baez, Doug Taylor, and Stephen Padgette for support throughout this work.

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Authors and Affiliations

  1. Monsanto Company, 700 Chesterfield Village Parkway North, St. Louis, MO 63198

    Jeffrey M. Staub, Peter T. J. Hajdukiewicz, Priscilla Hunter, Narender Nehra, Michael Schlittler, James A. Carroll, Dannette Ward & Guangning Ye

  2. Integrated Protein Technologies, Agracetus Campus, Monsanto Company, 8520 University Green, Middleton, 53562, WI

    Bradley Garcia, Julie Graves, Vikram Paradkar, Lori Spatola & Douglas A. Russell

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  1. Jeffrey M. Staub
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  2. Bradley Garcia
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Correspondence to Jeffrey M. Staub.

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Staub, J., Garcia, B., Graves, J. et al. High-yield production of a human therapeutic protein in tobacco chloroplasts . Nat Biotechnol 18, 333–338 (2000). https://doi.org/10.1038/73796

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