title: Biochemical and functional characterization of anti-HIV antibody-ELP fusion proteins from transgenic plants.
abstract: The stability and recovery of recombinant proteins expressed in plants are improved by fusion to elastin-like peptides (ELPs). In order to test the suitability of ELP for the production of pharmaceutical proteins, transgenic plants were created that individually expressed the light and heavy chains of the broadly neutralizing anti-human immunodeficiency virus type 1 (anti-HIV-1) monoclonal antibody 2F5, which is being evaluated as a microbicide component. The antibody chains were expressed both with and without a C-terminal ELP fusion. Crossing these plants in all combinations resulted in transgenic lines producing the full antibody in four formats, with ELP on either the light or heavy chains, on both or on neither. Characterization of the affinity-purified antibodies by surface plasmon resonance spectroscopy showed that the kinetic binding parameters were identical to those of a Chinese hamster ovary (CHO) cell counterpart lacking ELP. N-Glycan analysis showed that all four derivatives contained predominantly oligo-mannose-type N-glycans and that the ELP fusions had no significant effect on N-glycan structure. It was concluded that ELP fusion to the light chain, heavy chain or both chains of a plant-derived antibody had no adverse affects on protein quality, but had a positive impact on the yield. ELP fusions do not interfere with folding, assembly, trafficking in the secretory pathway or post-translational modification, but enhance stability whilst at the same time simplifying recovery.
authors.affiliation: Leibniz Institute of Plant Genetics and Crop Plant Research (IPK), Corrensstrasse 3, D-06466 Gatersleben, Germany.
authors.forename: Doreen M
authors.initials: DM
authors.lastname: Floss
authors.forename: Markus
authors.initials: M
authors.lastname: Sack
authors.forename: Johannes
authors.initials: J
authors.lastname: Stadlmann
authors.forename: Thomas
authors.initials: T
authors.lastname: Rademacher
authors.forename: Jürgen
authors.initials: J
authors.lastname: Scheller
authors.forename: Eva
authors.initials: E
authors.lastname: Stöger
authors.forename: Rainer
authors.initials: R
authors.lastname: Fischer
authors.forename: Udo
authors.initials: U
authors.lastname: Conrad
mesh_terms: D004549:Elastin / Q000235:genetics / Q000378:metabolism*; D015870:Gene Expression; D018506:Gene Expression Regulation, Plant; D015483:HIV Antibodies / Q000235:genetics / Q000276:immunology* / Q000378:metabolism*; D018515:Plant Leaves / Q000378:metabolism; D030821:Plants, Genetically Modified; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D014026:Nicotiana / Q000235:genetics / Q000378:metabolism*; D019076:Transgenes / Q000235:genetics*
chemical_list: D015483:HIV Antibodies; D011993:Recombinant Fusion Proteins; D004549:Elastin