title: Use of transgenic oryzacystatin-I-expressing plants enhances recombinant protein production.
abstract: Plants are an effective and inexpensive host for the production of commercially interesting heterologous recombinant proteins. The Escherichia coli-derived glutathione reductase was transiently expressed as a recombinant model protein in the cytosol of tobacco plants using the technique of leaf agro-infiltration. Proteolytic cysteine protease activity progressively increased over time when glutathione reductase accumulated in leaves. Application of cysteine protease promoter-GUS fusions in transgenic tobacco identified a cysteine protease NtCP2 expressed in mature leaves and being stress responsive to be expressed as a consequence of agro-infiltration. Transgenic tobacco plants constitutively expressing the rice cysteine protease inhibitor oryzacystatin-I had significantly lower cysteine protease activity when compared to non-transgenic tobacco plants. Lower cysteine protease activity in transgenic plants was directly related to higher glutathione reductase activity and also higher glutathione reductase amounts in transgenic plants. Overall, our work has demonstrated as a novel aspect that transgenic tobacco plants constitutively expressing an exogenous cysteine protease inhibitor have the potential for producing more recombinant protein which is very likely due to the reduced activity of endogenous cysteine protease.
authors.affiliation: Department of Plant Science, Forestry and Agricultural Biotechnology Institute, University of Pretoria, Pretoria 0002, South Africa.
authors.forename: P
authors.initials: P
authors.lastname: Pillay
authors.forename: T
authors.initials: T
authors.lastname: Kibido
authors.forename: M
authors.initials: M
authors.lastname: du Plessis
authors.forename: C
authors.initials: C
authors.lastname: van der Vyver
authors.forename: G
authors.initials: G
authors.lastname: Beyene
authors.forename: B J
authors.initials: BJ
authors.lastname: Vorster
authors.forename: K J
authors.initials: KJ
authors.lastname: Kunert
authors.forename: U
authors.initials: U
authors.lastname: Schlüter
mesh_terms: D057056:Cysteine Proteases / Q000378:metabolism; D015853:Cysteine Proteinase Inhibitors / Q000235:genetics / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D005818:Genetic Engineering / Q000379:methods*; D005980:Glutathione Reductase / Q000235:genetics* / Q000378:metabolism; D012275:Oryza / Q000235:genetics*; D010940:Plant Proteins / Q000235:genetics* / Q000378:metabolism; D030821:Plants, Genetically Modified / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D055333:Salivary Cystatins / Q000235:genetics* / Q000378:metabolism; D014026:Nicotiana / Q000235:genetics / Q000378:metabolism*; D015854:Up-Regulation
chemical_list: D015853:Cysteine Proteinase Inhibitors; D010940:Plant Proteins; D011994:Recombinant Proteins; D055333:Salivary Cystatins; D005980:Glutathione Reductase; D057056:Cysteine Proteases