title: Design and establishment of a vector system that enables production of multifusion proteins and easy purification by a two-step affinity chromatography approach.
abstract: The LE (LguI/Eco81I)-cloning procedure allows a step-wise, directional fusion of multiple DNA-fragments into a vector by utilizing two restriction enzymes generating identical non-palindromic overhangs. This strategy was applied to produce heat-stable cellulase-fusion proteins containing up to five single moieties. Terminal affinity tags enable efficient purification using a simple two-step approach.
authors.affiliation: Technische Universität Hamburg-Harburg, Institut für Technische Mikrobiologie, Kasernenstr. 12, D-21073 Hamburg, Germany.
authors.forename: Tabea
authors.initials: T
authors.lastname: Marquardt
authors.affiliation: Technische Universität Hamburg-Harburg, Institut für Technische Mikrobiologie, Kasernenstr. 12, D-21073 Hamburg, Germany.
authors.forename: Amélie
authors.initials: A
authors.lastname: von der Heyde
authors.affiliation: Technische Universität Hamburg-Harburg, Institut für Technische Mikrobiologie, Kasernenstr. 12, D-21073 Hamburg, Germany. Electronic address: skander.elleuche@tuhh.de.
authors.forename: Skander
authors.initials: S
authors.lastname: Elleuche
mesh_terms: D002482:Cellulose / Q000235:genetics / Q000302:isolation & purification; D002846:Chromatography, Affinity / Q000379:methods*; D015202:Protein Engineering / Q000379:methods*; D011993:Recombinant Fusion Proteins / Q000235:genetics* / Q000302:isolation & purification*
chemical_list: D011993:Recombinant Fusion Proteins; D002482:Cellulose
keywords: Gene fusion; Heat-stable cellulases; Multiple fusion approach; Two-step affinity chromatography; Type IIS restriction enzyme