title: Improving scFv antibody expression levels in the plant cytosol.
abstract: Expression of single-chain antibody fragments (scFvs) in the plant cytosol is often cumbersome. It was unexpectedly shown that addition at the C-terminus of the ER retention signal KDEL resulted in significantly improved expression levels. In this report the cytosolic location of the scFv-CK was confirmed, excluding possible mistranslocation to other subcellular compartments. It was shown that expression of several other scFvs was also improved in tobacco protoplasts. In addition expression was improved in transgenic potato. Changing from KDEL to KDEI did not affect the enhanced protein expression level. Addition of the KDEL motif is a simple and straightforward tool to stabilize in planta cytosolic expression of many scFvs.
authors.affiliation: Department of Nematology, Wageningen Agricultural University, The Netherlands. sander.schouten@medew.nema.wau.nl
authors.forename: A
authors.initials: A
authors.lastname: Schouten
authors.forename: J
authors.initials: J
authors.lastname: Roosien
authors.forename: J M
authors.initials: JM
authors.lastname: de Boer
authors.forename: A
authors.initials: A
authors.lastname: Wilmink
authors.forename: M N
authors.initials: MN
authors.lastname: Rosso
authors.forename: D
authors.initials: D
authors.lastname: Bosch
authors.forename: W J
authors.initials: WJ
authors.lastname: Stiekema
authors.forename: F J
authors.initials: FJ
authors.lastname: Gommers
authors.forename: J
authors.initials: J
authors.lastname: Bakker
authors.forename: A
authors.initials: A
authors.lastname: Schots
mesh_terms: D002265:Carboxylic Ester Hydrolases / Q000276:immunology; D002460:Cell Line; D003001:Cloning, Molecular; D003600:Cytosol / Q000378:metabolism*; D004721:Endoplasmic Reticulum / Q000378:metabolism; D018506:Gene Expression Regulation, Plant; D006031:Glycosylation; D006825:Hybridomas; D007128:Immunoglobulin Fragments / Q000235:genetics*; D007135:Immunoglobulin Variable Region / Q000235:genetics* / Q000276:immunology; D008969:Molecular Sequence Data; D009842:Oligopeptides / Q000378:metabolism; D010455:Peptides / Q000737:chemistry / Q000378:metabolism; D030821:Plants, Genetically Modified; D010947:Plants, Toxic; D021382:Protein Sorting Signals / Q000737:chemistry / Q000378:metabolism; D011523:Protoplasts / Q000378:metabolism; D011198:Solanum tuberosum / Q000235:genetics* / Q000378:metabolism; D014026:Nicotiana / Q000235:genetics* / Q000378:metabolism / Q000648:ultrastructure; D014170:Transformation, Genetic
chemical_list: D007128:Immunoglobulin Fragments; D007135:Immunoglobulin Variable Region; D009842:Oligopeptides; D010455:Peptides; D021382:Protein Sorting Signals; C055966:immunoglobulin Fv; C073762:lysyl-aspartyl-glutamyl-leucine; D002265:Carboxylic Ester Hydrolases; C019952:cutinase