Head-to-tail fusions of camelid antibodies can be expressed in planta and bind in rumen fluid.
We have compared the accumulation of recombinant variable heavy-chain portions [VHH (variable heavy-chain antibody from camelids)] of camelid antibodies in a variety of subcellular compartments produced in planta. The VHH coding sequences were optimized for expression in thale cress (Arabidopsis thaliana) and placed individually or as fused tandem heterodimers in synthetic plant-organelle-targeting cassettes designed to target the protein to either the cytoplasm, ER (endoplasmic reticulum), protein storage vacuole or chloroplast. Accumulation of individual VHHs was only detected in plants transformed with the ER-targeting cassette, whereas accumulation of the tandem VHHs was detected for all cassettes and was the highest with the ER cassette [0.1-0.7% (w/w) of total soluble proteins]. The ability of the plant-produced tandem VHH to reduce TNFalpha (tumour necrosis factor alpha) cytotoxicity was found to be comparable with previously characterized recombinant VHHs. In vitro antigen binding and functional stability in rumen fluid were determined on both prokaryotically expressed and plant-expressed tandem VHHs. The plant-produced VHH did not appear to be any more stable in rumen fluid than other soluble plant proteins; however, it was able to bind equally well to the antigen in the presence or absence of rumen fluid.
Forage Biotechnology, AgResearch Grasslands, Palmerston North, New Zealand.
Somrutai
S
Winichayakul
Anton
A
Pernthaner
Richard
R
Scott
Ruth
R
Vlaming
Nick
N
Roberts
D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies / Q000737:chemistry / Q000235:genetics* / Q000276:immunology*; D017360:Arabidopsis / Q000166:cytology / Q000235:genetics* / Q000378:metabolism; D001483:Base Sequence; D001826:Body Fluids / Q000378:metabolism*; D002161:Camelids, New World / Q000235:genetics / Q000276:immunology*; D015870:Gene Expression; D007143:Immunoglobulin Heavy Chains / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D007135:Immunoglobulin Variable Region / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D008969:Molecular Sequence Data; D018515:Plant Leaves / Q000166:cytology / Q000235:genetics / Q000378:metabolism; D010940:Plant Proteins / Q000737:chemistry; D030821:Plants, Genetically Modified; D055550:Protein Stability; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics* / Q000276:immunology; D012417:Rumen / Q000378:metabolism*; D014409:Tumor Necrosis Factor-alpha / Q000276:immunology
D000906:Antibodies; D007143:Immunoglobulin Heavy Chains; D007135:Immunoglobulin Variable Region; D010940:Plant Proteins; D011993:Recombinant Fusion Proteins; D014409:Tumor Necrosis Factor-alpha