Arabidopsis thaliana Rubisco small subunit transit peptide increases the accumulation of Thermotoga maritima endoglucanase Cel5A in chloroplasts of transgenic tobacco plants.
Over the past decade various approaches have been used to increase the expression level of recombinant proteins in plants. One successful approach has been to target proteins to specific subcellular sites/compartments of plant cells, such as the chloroplast. In the study reported here, hyperthermostable endoglucanase Cel5A was targeted into the chloroplasts of tobacco plants via the N-terminal transit peptide of nuclear-encoded plastid proteins. The expression levels of Cel5A transgenic lines were then determined using three distinct transit peptides, namely, the light-harvesting chlorophyll a/b-binding protein (CAB), Rubisco small subunit (RS), and Rubisco activase (RA). RS:Cel5A transgenic lines produced highly stable active enzymes, and the protein accumulation of these transgenic lines was up to 5.2% of the total soluble protein in the crude leaf extract, remaining stable throughout the life cycle of the tobacco plant. Transmission election microscopy analysis showed that efficient targeting of Cel5A protein was under the control of the transit peptide.
Kim
Suyeon
S
Bio-energy Research Institute, Chonnam National University, Gwangju, 500-757, Republic of Korea.
Lee
Dae-Seok
DS
Choi
In Seong
IS
Ahn
Sung-Ju
SJ
Kim
Yong-Hwan
YH
Bae
Hyeun-Jong
HJ
D017360:Arabidopsis / Q000201:enzymology*; D002480:Cellulase / Q000378:metabolism* / Q000648:ultrastructure; D002736:Chloroplasts / Q000378:metabolism*; D017931:DNA Primers / Q000235:genetics; D015151:Immunoblotting; D007150:Immunohistochemistry; D046529:Microscopy, Electron, Transmission; D018515:Plant Leaves / Q000378:metabolism / Q000648:ultrastructure; D030821:Plants, Genetically Modified / Q000378:metabolism*; D010957:Plasmids / Q000235:genetics; D011994:Recombinant Proteins / Q000378:metabolism*; D012273:Ribulose-Bisphosphate Carboxylase / Q000378:metabolism*; D020124:Thermotoga maritima / Q000201:enzymology*; D014026:Nicotiana / Q000378:metabolism*; D014170:Transformation, Genetic
D017931:DNA Primers; D011994:Recombinant Proteins; C407024:endoglucanase Cel5A; D002480:Cellulase; D012273:Ribulose-Bisphosphate Carboxylase