Expression and subcellular targeting of human insulin-like growth factor binding protein-3 in transgenic tobacco plants.
Human insulin-like growth factor binding protein-3 (hIGFBP-3) is a multifunctional protein which has high affinity for insulin-like growth factor-I (IGF-I). It combines with IGF-I to form a tertiary complex in circulation, thus regulating the activity of IGF-I. Furthermore, recombinant hIGFBP-3 (rhIGFBP-3) has been found to negatively regulate cell proliferation and induce apoptosis. In this study, we have established an efficient plant bioreactor platform for mass production of rhIGFBP-3. Different expression constructs, driven by the seed-specific phaseolin promoter, were designed and transformed into tobacco plant via Agrobacterium. To enhance protein expression level, the signal peptide (SP) and the C-terminal tetrapeptide AFVY of phaseolin were used to direct rhIGFBP-3 to protein storage vacuole (PSV) in tobacco seed for stable accumulation. Western blot analysis showed that rhIGFBP-3 was successfully synthesized in transgenic tobacco seeds, with the highest protein expression of 800 mug/g dry weight. The localization of rhIGFBP-3 in PSV was also evident by confocal immunofluorescence microscopy. Our results indicated that protein sorting sequences could benefit the expression level of rhIGFBP-3 and it is feasible to use plant as "bio-factory" to produce therapeutic recombinant proteins in large quantity.
Cheung
Stanley C K
SC
Department of Medicine and Therapeutics, Prince of Wales Hospital, The Chinese University of Hong Kong, Shatin, Hong Kong SAR, China.
Sun
Samuel S M
SS
Chan
Juliana C N
JC
Tong
Peter C Y
PC
D018745:Genome, Plant; D006801:Humans; D018972:Insulin-Like Growth Factor Binding Protein 3; D018969:Insulin-Like Growth Factor Binding Proteins / Q000235:genetics* / Q000378:metabolism; D030821:Plants, Genetically Modified / Q000235:genetics; D021382:Protein Sorting Signals; D021381:Protein Transport; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D012231:Rhizobium / Q000235:genetics; D014026:Nicotiana / Q000235:genetics* / Q000378:metabolism; D014170:Transformation, Genetic
C515497:IGFBP3 protein, human; D018972:Insulin-Like Growth Factor Binding Protein 3; D018969:Insulin-Like Growth Factor Binding Proteins; D021382:Protein Sorting Signals; D011994:Recombinant Proteins