title: Companion Protease Inhibitors for the In Situ Protection of Recombinant Proteins in Plants.
abstract: We previously described a procedure for the use of plant protease inhibitors as "companion" accessory proteins to prevent unwanted proteolysis of clinically useful recombinant proteins in leaf crude protein extracts (Benchabane et al. Methods Mol Biol 483:265-273, 2009). Here we describe the use of these inhibitors for the protection of recombinant proteins in planta, before their extraction from leaf tissues. A procedure is first described involving inhibitors co-expressed along-and co-migrating-with the protein of interest in host plant cells. An alternative, single transgene scheme is then described involving translational fusions of the recombinant protein and companion inhibitor. These approaches may allow for a significant improvement of protein steady-state levels in leaves, comparable to yield improvements observed with protease-deficient strains of less complex protein expression hosts such as E. coli or yeasts.
authors.affiliation: Centre de Recherche et d'Innovation sur les Végétaux, Université Laval, Québec, QC, Canada.
authors.forename: Stéphanie
authors.initials: S
authors.lastname: Robert
authors.affiliation: Centre de Recherche et d'Innovation sur les Végétaux, Université Laval, Québec, QC, Canada.
authors.forename: Philippe V
authors.initials: PV
authors.lastname: Jutras
authors.affiliation: Centre de Recherche et d'Innovation sur les Végétaux, Université Laval, Québec, QC, Canada.
authors.forename: Moustafa
authors.initials: M
authors.lastname: Khalf
authors.affiliation: Medicago Inc., Québec, QC, Canada.
authors.forename: Marc-André
authors.initials: MA
authors.lastname: D'Aoust
authors.affiliation: Centre de Recherche et d'Innovation sur les Végétaux, Université Laval, Québec, QC, Canada.
authors.forename: Marie-Claire
authors.initials: MC
authors.lastname: Goulet
authors.affiliation: Centre de Recherche et d'Innovation sur les Végétaux, Université Laval, Québec, QC, Canada.|Australian Institute for Bioengineering and Nanotechnology, The University of Queensland, St Lucia, QLD, Australia.
authors.forename: Frank
authors.initials: F
authors.lastname: Sainsbury
authors.affiliation: Centre de Recherche et d'Innovation sur les Végétaux, Université Laval, Québec, QC, Canada. dominique.michaud@fsaa.ulaval.ca.
authors.forename: Dominique
authors.initials: D
authors.lastname: Michaud
mesh_terms: D005818:Genetic Engineering / Q000379:methods*; D030821:Plants, Genetically Modified; D011480:Protease Inhibitors / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism*; D014026:Nicotiana / Q000235:genetics* / Q000378:metabolism; D019076:Transgenes
chemical_list: D011480:Protease Inhibitors; D011994:Recombinant Proteins
keywords: Clinically useful recombinant proteins; Companion protease inhibitors; Heterologous protein expression; Protein stabilization; Recombinant protein degradation