Expression of non-toxic mutant of Escherichia coli heat-labile enterotoxin in tobacco chloroplasts.
Chloroplast transformation systems offer unique advantages in biotechnology, including high level of foreign gene expression, maternal inheritance, and polycistronic expression. We studied chloroplast expression of LTK63 (change Ser-->Lys at position 63 in the A subunit) which is the mutant of Escherichia coli heat-labile toxin. LTK63 is devoid of any toxic activity, but still retains its mucosal adjuvanticity. The LTK63 was cloned into chloroplast targeting vector and transformed to tobacco chloroplasts by particle bombardment. PCR and Southern blot analyses confirmed stable homologous recombination of the LTK63 gene into the chloroplast genome. The amount of LTK63 protein detected in tobacco chloroplasts was approximately 3.7% of the total soluble protein. The GM1-ganglioside binding assay confirmed that chloroplast-synthesized LTB of LTK63 binds to the intestinal membrane GM1-ganglioside receptor. Thus, the expression of LTK63 in chloroplasts provides a potential route toward the development of a plant-based edible vaccine for high expression system and environmentally friendly approach.
Kang
Tae-Jin
TJ
Institute of Basic Science, Chonbuk National University, Jeonju 561-756, South Korea.
Han
So-Chon
SC
Kim
Mi-Young
MY
Kim
Young-Sook
YS
Yang
Moon-Sik
MS
D001427:Bacterial Toxins / Q000235:genetics* / Q000378:metabolism; D002736:Chloroplasts / Q000378:metabolism*; D004768:Enterotoxins / Q000235:genetics* / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000235:genetics* / Q000378:metabolism; D005822:Genetic Vectors; D015151:Immunoblotting; D020161:Physical Chromosome Mapping; D030821:Plants, Genetically Modified; D010957:Plasmids; D011485:Protein Binding; D011956:Receptors, Cell Surface / Q000378:metabolism; D014026:Nicotiana / Q000235:genetics*; D014170:Transformation, Genetic
D001427:Bacterial Toxins; D004768:Enterotoxins; D029968:Escherichia coli Proteins; D011956:Receptors, Cell Surface; C034993:ganglioside receptor; C027538:heat-labile enterotoxin, E coli