Expression of the B subunit of E. coli heat-labile enterotoxin in the chloroplasts of plants and its characterization.
Transgenic chloroplasts have become attractive systems for heterologous gene expressions because of unique advantages. Here, we report a feasibility study for producing the nontoxic B subunit of Escherichia coli heat-labile enterotoxin (LTB) via chloroplast transformation of tobacco. Stable site-specific integration of the LTB gene into chloroplast genome was confirmed by PCR and genomic Southern blot analysis in transformed plants. Immunoblot analysis indicated that plant-derived LTB protein was oligomeric, and dissociated after boiling. Pentameric LTB molecules were the dominant molecular species in LTB isolated from transgenic tobacco leaf tissues. The amount of LTB protein detected in transplastomic tobacco leaf was approximately 2.5% of the total soluble plant protein, approximately 250-fold higher than in plants generated via nuclear transformation. The GM1-ELISA binding assay indicated that chloroplast-synthesized LTB protein bound to GM1-ganglioside receptors. LTB protein with biochemical properties identical to native LTB protein in the chloroplast of edible plants opens the way for inexpensive, safe, and effective plant-based edible vaccines for humans and animals.
Kang
Tae-Jin
TJ
Basic Sciences Research Institute, Chonbuk National University, Jeonju 561-756, South Korea.
Loc
Nguyen-Hoang
NH
Jang
Mi-Ok
MO
Jang
Yong-Suk
YS
Kim
Young-Sook
YS
Seo
Jo-Eun
JE
Yang
Moon-Sik
MS
D001427:Bacterial Toxins / Q000096:biosynthesis* / Q000737:chemistry; D002736:Chloroplasts / Q000235:genetics / Q000378:metabolism*; D004768:Enterotoxins / Q000096:biosynthesis* / Q000737:chemistry; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D029968:Escherichia coli Proteins; D018014:Gene Transfer Techniques; D020161:Physical Chromosome Mapping; D030821:Plants, Genetically Modified; D010947:Plants, Toxic; D018087:Plastids / Q000235:genetics; D014026:Nicotiana / Q000235:genetics*; D014170:Transformation, Genetic
D001427:Bacterial Toxins; D004768:Enterotoxins; D029968:Escherichia coli Proteins; C027538:heat-labile enterotoxin, E coli