Molecular design of photosynthesis-elevated chloroplasts for mass accumulation of a foreign protein.
In order to increase production of a useful protein by the chloroplast transformation technique, it seems to be necessary to determine the 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 200-260 mg ml(-1) of green fluorescent protein in the stroma without any inhibition of gas exchange under various light intensity and growth conditions. 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- to 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 <1 mg ml(-1) of the FBP/SBPase protein in chloroplasts and were calculated to represent <1% of the protein accumulated via chloroplast transformation. Consequently, >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.
Yabuta
Yukinori
Y
Department of Advanced Bioscience, Faculty of Agriculture, Kinki University, 3327-204 Nakamachi, Nara 631-8505, Japan.
Tamoi
Masahiro
M
Yamamoto
Kumiko
K
Tomizawa
Ken-Ichi
K
Yokota
Akiho
A
Shigeoka
Shigeru
S
D002736:Chloroplasts / Q000378:metabolism*; D006597:Fructose-Bisphosphatase / Q000235:genetics / Q000378:metabolism*; D018506:Gene Expression Regulation, Plant; D005818:Genetic Engineering / Q000379:methods*; D010744:Phosphoric Monoester Hydrolases / Q000235:genetics / Q000378:metabolism*; D010788:Photosynthesis / Q000502:physiology*; D018515:Plant Leaves / Q000201:enzymology; D030821:Plants, Genetically Modified; D014026:Nicotiana / Q000201:enzymology / Q000235:genetics*
D006597:Fructose-Bisphosphatase; D010744:Phosphoric Monoester Hydrolases; C022627:sedoheptulose-bisphosphatase