title: Tandem SUMO fusion vectors for improving soluble protein expression and purification.
abstract: Availability of highly purified proteins in quantity is crucial for detailed biochemical and structural investigations. Fusion tags are versatile tools to facilitate efficient protein purification and to improve soluble overexpression of proteins. Various purification and fusion tags have been widely used for overexpression in Escherichia coli. However, these tags might interfere with biological functions and/or structural investigations of the protein of interest. Therefore, an additional purification step to remove fusion tags by proteolytic digestion might be required. Here, we describe a set of new vectors in which yeast SUMO (SMT3) was used as the highly specific recognition sequence of ubiquitin-like protease 1, together with other commonly used solubility enhancing proteins, such as glutathione S-transferase, maltose binding protein, thioredoxin and trigger factor for optimizing soluble expression of protein of interest. This tandem SUMO (T-SUMO) fusion system was tested for soluble expression of the C-terminal domain of TonB from different organisms and for the antiviral protein scytovirin.
authors: Research Program in Structural Biology and Biophysics, Institute of Biotechnology, University of Helsinki, P.O. Box 65, Helsinki FIN-00014, Finland.
authors: Fernando
authors: F
authors: Guerrero
authors: Research Program in Structural Biology and Biophysics, Institute of Biotechnology, University of Helsinki, P.O. Box 65, Helsinki FIN-00014, Finland.
authors: Annika
authors: A
authors: Ciragan
authors: Research Program in Structural Biology and Biophysics, Institute of Biotechnology, University of Helsinki, P.O. Box 65, Helsinki FIN-00014, Finland. Electronic address: hideo.iwai@helsinki.fi.
authors: Hideo
authors: H
authors: Iwaï
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D001483:Base Sequence; D002352:Carrier Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D003001:Cloning, Molecular / Q000379:methods*; D000458:Cyanobacteria / Q000737:chemistry / Q000235:genetics; D003546:Cysteine Endopeptidases / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification; D004926:Escherichia coli / Q000737:chemistry / Q000235:genetics; D005822:Genetic Vectors / Q000737:chemistry / Q000235:genetics*; D016480:Helicobacter pylori / Q000737:chemistry / Q000235:genetics; D037102:Lectins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D059748:Proteolysis; D011549:Pseudomonas / Q000737:chemistry / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D012441:Saccharomyces cerevisiae / Q000737:chemistry / Q000235:genetics*; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification; D025841:Small Ubiquitin-Related Modifier Proteins / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification; D012995:Solubility
chemical_list: D001426:Bacterial Proteins; D002352:Carrier Proteins; D037102:Lectins; D008565:Membrane Proteins; D011993:Recombinant Fusion Proteins; C099362:SMT3 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D025841:Small Ubiquitin-Related Modifier Proteins; C474342:scytovirin protein, S varium; C057616:tonB protein, Bacteria; D003546:Cysteine Endopeptidases; C118083:Ulp1 protease
keywords: Fusion proteins; Protein ligation; SUMO system; Scytovirin; Solubility enhancement tag; TonB; Ulp1 protease