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Improved stearate phenotype in transgenic canola expressing a modified acyl-acyl carrier protein thioesterase

Nature Biotechnology volume 17, pages 593–597 (1999)Cite this article

Abstract

The engineering of crops for selected fatty acid production is one of the major goals of plant biotechnology. The Garm FatA1, an acyl-acyl carrier protein (ACP) thioesterase isolated from Garcinia mangostana, generates an elevated stearate (18:0) phenotype in transgenic Brassica plants. By site-directed mutagenesis, we generated seven mutants that showed up to a 13-fold increase in specific enzyme activity toward 18:0-ACP in vitro. The seed-specific expression of mutant S111A/V193A in Brassica plants results in transgenic plants that accumulate 55–68% more stearate than plants expressing the wild-type enzyme. Our results demonstrate that a thioesterase can be engineered to increase specific activity and that its improved function demonstrated in vitro is retained in vivo.

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Figure 1: Amino acid alignment of the six positions where the residues in Garm FatA1 differ from what otherwise are completely conserved in all other known FatA TEs.
Figure 2: Relative enzyme activities of the wild-type and mutant Garm FatA1.
Figure 3: Scheme of the tDNA components of the binary vectors for plant transformation.
Figure 4: Stearate accumulation of Brassica seeds expressing the wild-type and mutant Garm FatA1.

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Acknowledgements

We are grateful to J. Kridl and D. Hawkins of Calgene for providing pGCN5255 and pCGN5266, and for their helpful suggestions throughout the study. We thank our colleagues T. Hayes, B. Reed, J. Turner, S. Radke, and B. Schreckengost for their technical support. Our thanks also go to V. Knauf, D. Facciotti, M. Lassner, and J. Metz of Calgene; as well as R. Glaeser and S. Rouhani of University of California at Berkeley; and E. Carlson of University of California at Davis, for their critical reading of the manuscript.

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  1. Marc T. Facciotti

    Present address: The Graduate Group in Biophysics, University of California at Berkeley, Berkeley, CA, 94720

  2. Ling Yuan

    Present address: Maxygen, Redwood City, CA, 94063

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  1. Calgene, LLC, Davis, 95616, CA

    Paul B. Bertain

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  1. Marc T. Facciotti
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  2. Paul B. Bertain
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  3. Ling Yuan
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Correspondence to Ling Yuan.

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Facciotti, M., Bertain, P. & Yuan, L. Improved stearate phenotype in transgenic canola expressing a modified acyl-acyl carrier protein thioesterase. Nat Biotechnol 17, 593–597 (1999). https://doi.org/10.1038/9909

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