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Plant and Cell Physiology Advance Access published online on January 25, 2008
Plant and Cell Physiology, doi:10.1093/pcp/pcn014
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© The Author 2008. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For Permissions, please e-mail: journals.permissions@oxfordjournals.org

Molecular designing of photosynthesis-elevated chloroplasts for mass accumulation of a foreign protein

Yukinori Yabuta1,4,5, Masahiro Tamoi1,5, Kumiko Yamamoto1, Ken-ichi Tomizawa2, Akiho Yokota3 and Shigeru Shigeoka1,*

1Department of Advanced Bioscience, Faculty of Agriculture, Kinki University, 3327-204 Nakamachi, Nara 631-8505, Japan
2Research Institute of Innovative Technology for the Earth, 9-2 Kizugawadai, Kizu-cho, Soraku-gun, Kyoto 619-0292, Japan
3Graduate School of Biological Sciences, Nara Institute of Science and Technology, 8916-5 Takayama, Ikoma, Nara 630-0101, Japan

Corresponding author: Prof. Shigeru Shigeokat. Department of Advanced Bioscience, Faculty of Agriculture, Kinki University, 3327-204 Nakamachi, Nara 631-8505, Japan Tel: +81-742-43-8083, FAX: +81-742-43-8976, E-mail: shigeoka{at}nara.kindai.ac.jp


  Abstract

In order to increase production of a useful protein by chloroplast transformation technique, it seems to be necessary to determine upper limit for the accumulation of a biologically active foreign protein in chloroplasts and then improve photosynthetic capacity and plant productivity. Here we show that the stromal fractions of tobacco chloroplasts could accommodate an additional 2007sim;260 mg ml–1 of green fluorescence protein in the stroma without any inhibition of gas exchange under various light intensity and growth. The minimum amount of fructose-1,6-/sedoheptulose-1,7-bisphosphatase (FBP/SBPase) limiting photosynthesis was then calculated. Analyses of the photosynthetic parameters and the metabolites of transformants into which FBP/SBPase was introduced with various types of promoter (PpsbA, Prrn, Prps2 and Prps12) indicated that a 2~3-fold increase in levels of FBPase and SBPase activity is sufficient to increase the final amount of dry matter by up to 1.8-fold relative to the wild-type plants. Their increases were equivalent to an increase of less than 1 mg ml–1 of the FBP/SBPase protein in chloroplasts and were calculated to represent less than 1% of the protein accumulated via chloroplast transformation. Consequently, more than 99% of the additional 200~260 mg ml–1 of protein expressed in the chloroplasts could be used for the production of useful proteins in the photosynthesis-elevated transplastomic plants having FBP/SBPase.

Keywords: plastid transformation - photosynthesis - fructose-1,6-/sedoheptulose-1 - 7-bisphosphatase - the Calvin cycle

4Present address: School of Agricultural, Biological, and Environmental Sciences, Faculty of Agriculture, Tottori University, Tottori 680-8553, Japan

5These authors contributed equally to this work.

(Received December 4, 2007; Accepted January 22, 2008)
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