High-level expression of a human β-site APP cleaving enzyme in transgenic tobacco chloroplasts and its immunogenicity in mice.
Plastid transformation has to date been applied to the expression of heterologous genes involved in agronomic traits and to the production of useful recombinant proteins. Here, we report a feasibility study for producing the human β-site APP cleaving enzyme (BACE) via transformation of tobacco chloroplasts. Stable integration of human BACE into the plastome was confirmed by PCR. Genomic Southern blot analysis detected the presence of the tobacco aadA and human BACE genes between trnI and trnA in the plastome. Northern blot analysis revealed that the aadA and BACE genes were both properly transcribed into a dicistronic transcriptional unit. Human BACE protein expression in transplastomic tobacco was determined by western blot analysis. ELISA analysis revealed that, based on a dilution series of E. coli-derived BACE as a standard, transplastomic lines accumulated BACE to levels of 2.0% of total soluble proteins. When mice were gavaged with the transplastomic tobacco extracts, they showed an immune response against the BACE antigen. The successful production of plastid-based BACE protein has the potential for developing a plant-based vaccine against Alzheimer disease.
Youm
Jung Won
JW
Plant Systems Engineering Research Center, KRIBB, Daejeon, 305-806, Korea.
Jeon
Jae Heung
JH
Kim
Hee
H
Min
Sung Ran
SR
Kim
Mi Sun
MS
Joung
Hyouk
H
Jeong
Won Joong
WJ
Kim
Hyun Soon
HS
D000544:Alzheimer Disease / Q000201:enzymology / Q000276:immunology / Q000517:prevention & control; D053829:Amyloid Precursor Protein Secretases / Q000096:biosynthesis / Q000235:genetics* / Q000276:immunology*; D000818:Animals; D016282:Aspartic Acid Endopeptidases / Q000096:biosynthesis / Q000235:genetics* / Q000276:immunology*; D001483:Base Sequence; D002736:Chloroplasts / Q000201:enzymology / Q000235:genetics; D017931:DNA Primers / Q000235:genetics; D005260:Female; D015870:Gene Expression; D017343:Genes, Plant; D006801:Humans; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D030821:Plants, Genetically Modified; D015202:Protein Engineering; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology; D014026:Nicotiana / Q000201:enzymology* / Q000235:genetics*; D014614:Vaccines, Synthetic / Q000235:genetics / Q000276:immunology
D017931:DNA Primers; D011994:Recombinant Proteins; D014614:Vaccines, Synthetic; D053829:Amyloid Precursor Protein Secretases; D016282:Aspartic Acid Endopeptidases; C509280:BACE1 protein, human