Production of Human Serum Albumin in Chloroplast-Transformed Tobacco Plants
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Human serum albumin (HSA) is the most abundant proteinin plasma and is the most often used intravenous protein inmany human therapies. However, HSA is currently extractedonly from plasma because commercially feasible recombinantexpression systems are not available. This studyattempted to develop an efficient system for recombinantHSA production by chloroplast transformation of tobacco. AHSA cDNA was isolated from a cDNA library constructedwith human liver tissue. Chloroplast transformation vectorswere constructed by introducing various regulatory elementsto HSA regulatory sequences. Vectors were delivered byparticle bombardment into leaf explants and chloroplasttransformedplants were subsequently regenerated intowhole plants. Southern blot analysis confirmed that the HSAcDNA was incorporated between rps12 and orf70B of thechloroplast genome as designed. Western blot analysisrevealed that hyper-expression and increasing the stabilityof HSA were achieved by modification of the regulatorysequences using the psbA5'UTRs in combination with elementsof the 14 N-terminal amino acids of the GFP and theFLAG tag. However, only plants transformed with the vectorcontaining all of these elements were able to accumulateHSA.
Abstract
Human serum albumin (HSA) is the most abundant proteinin plasma and is the most often used intravenous protein inmany human therapies. However, HSA is currently extractedonly from plasma because commercially feasible recombinantexpression systems are not available. This studyattempted to develop an efficient system for recombinantHSA production by chloroplast transformation of tobacco. AHSA cDNA was isolated from a cDNA library constructedwith human liver tissue. Chloroplast transformation vectorswere constructed by introducing various regulatory elementsto HSA regulatory sequences. Vectors were delivered byparticle bombardment into leaf explants and chloroplasttransformedplants were subsequently regenerated intowhole plants. Southern blot analysis confirmed that the HSAcDNA was incorporated between rps12 and orf70B of thechloroplast genome as designed. Western blot analysisrevealed that hyper-expression and increasing the stabilityof HSA were achieved by modification of the regulatorysequences using the psbA5'UTRs in combination with elementsof the 14 N-terminal amino acids of the GFP and theFLAG tag. However, only plants transformed with the vectorcontaining all of these elements were able to accumulateHSA.