High-level expression of Camelid nanobodies in Nicotiana benthamiana.
Nanobodies (or VHHs) are single-domain antigen-binding fragments derived from Camelid heavy chain-only antibodies. Their small size, monomeric behaviour, high stability and solubility, and ability to bind epitopes not accessible to conventional antibodies make them especially suitable for many therapeutic and biotechnological applications. Here we describe high-level expression, in Nicotiana benthamiana, of three versions of an anti-hen egg white lysozyme (HEWL) nanobody which include the original VHH from an immunized library (cAbLys3), a codon-optimized derivative, and a codon-optimized hybrid nanobody comprising the CDRs of cAbLys3 grafted onto an alternative 'universal' nanobody framework. His6- and StrepII-tagged derivatives of each nanobody were targeted for accumulation in the cytoplasm, chloroplast and apoplast using different pre-sequences. When targeted to the apoplast, intact functional nanobodies accumulated at an exceptionally high level (up to 30% total leaf protein), demonstrating the great potential of plants as a nanobody production system.
Plant Molecular Genetics Laboratory, Smurfit Institute of Genetics, Trinity College, Dublin 2, Ireland.
Yi-Hui Audrey
YH
Teh
Tony A
TA
Kavanagh
D016960:Agrobacterium tumefaciens / Q000235:genetics; D000595:Amino Acid Sequence; D000818:Animals; D000910:Antibodies, Heterophile / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D000917:Antibody Formation / Q000235:genetics / Q000502:physiology; D001483:Base Sequence; D002161:Camelids, New World / Q000276:immunology*; D002645:Chickens / Q000235:genetics; D003001:Cloning, Molecular; D007143:Immunoglobulin Heavy Chains / Q000378:metabolism; D008969:Molecular Sequence Data; D009113:Muramidase / Q000235:genetics / Q000378:metabolism; D030821:Plants, Genetically Modified; D012689:Sequence Homology, Nucleic Acid; D014026:Nicotiana / Q000235:genetics / Q000378:metabolism*
D000910:Antibodies, Heterophile; D007143:Immunoglobulin Heavy Chains; D009113:Muramidase