title: DSF Guided Refolding As A Novel Method Of Protein Production.
abstract: The Anfinsen hypothesis, the demonstration of which led to the Nobel prize in Chemistry, posits that all information required to determine a proteins' three dimensional structure is contained within its amino acid sequence. This suggests that it should be possible, in theory, to fold any protein in vitro. In practice, however, protein production by refolding is challenging because suitable refolding conditions must be empirically determined for each protein and can be painstaking. Here we demonstrate, using a variety of proteins, that differential scanning fluorimetry (DSF) can be used to determine and optimize conditions that favor proper protein folding in a rapid and high-throughput fashion. The resulting method, which we deem DSF guided refolding (DGR), thus enables the production of aggregation-prone and disulfide-containing proteins by refolding from E. coli inclusion bodies, which would not normally be amenable to production in bacteria.
authors.affiliation: Diabetes and Metabolic Disease Program, Houston Methodist Research Institute, Houston, Texas, USA.
authors.forename: Amadeo B
authors.initials: AB
authors.lastname: Biter
authors.affiliation: Department of Biomedical Engineering, Johns Hopkins University, Baltimore, Maryland, USA.
authors.forename: Andres H
authors.initials: AH
authors.lastname: de la Peña
authors.affiliation: Department of BioSciences-Biochemistry and Cell Biology, Rice University, Houston, Texas, USA.
authors.forename: Roopa
authors.initials: R
authors.lastname: Thapar
authors.affiliation: Diabetes and Metabolic Disease Program, Houston Methodist Research Institute, Houston, Texas, USA.
authors.forename: Jean Z
authors.initials: JZ
authors.lastname: Lin
authors.affiliation: Department of Molecular and Cellular Biology, Baylor College of Medicine, Houston, Texas, USA.
authors.forename: Kevin J
authors.initials: KJ
authors.lastname: Phillips
mesh_terms: D000595:Amino Acid Sequence / Q000235:genetics; D000596:Amino Acids / Q000737:chemistry* / Q000235:genetics; D004220:Disulfides / Q000737:chemistry; D004926:Escherichia coli / Q000737:chemistry* / Q000235:genetics; D005470:Fluorometry; D002479:Inclusion Bodies / Q000737:chemistry; D017510:Protein Folding; D011506:Proteins / Q000737:chemistry* / Q000235:genetics
chemical_list: D000596:Amino Acids; D004220:Disulfides; D011506:Proteins