title: Rapid matrix-assisted refolding of histidine-tagged proteins.
abstract: The formation of inclusion bodies (IBs)--amorphous aggregates of misfolded insoluble protein--during recombinant protein expression, is still one of the biggest bottlenecks in protein science. We have developed and analyzed a rapid parallel approach for matrix-assisted refolding of recombinant His(6)-tagged proteins. Efficiencies of matrix-assisted refolding were screened in a 96-well format. The developed methodology allowed the efficient refolding of five different test proteins, including monomeric and oligomeric proteins. Compared to refolding in-solution, the matrix-assisted refolding strategy proved equal or better for all five proteins tested. Interestingly, specifically oligomeric proteins displayed significantly higher levels of refolding compared to refolding in-solution. Mechanistically, matrix-assisted folding seems to differ from folding in-solution, as the reaction proceeds more rapidly and shows a remarkably different concentration dependence--it allows refolding at up to 1000-fold higher protein concentration than folding in-solution.
authors.affiliation: Munich Center for Integrated Protein Science and Chemistry Department, Technische Universität München, 85747 Garching, Germany.
authors.forename: Tetyana
authors.initials: T
authors.lastname: Dashivets
authors.forename: Nichole
authors.initials: N
authors.lastname: Wood
authors.forename: Christoph
authors.initials: C
authors.lastname: Hergersberg
authors.forename: Johannes
authors.initials: J
authors.lastname: Buchner
authors.forename: Martin
authors.initials: M
authors.lastname: Haslbeck
mesh_terms: D000818:Animals; D001681:Biological Assay / Q000379:methods*; D002021:Buffers; D049452:Green Fluorescent Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D006639:Histidine / Q000378:metabolism*; D002479:Inclusion Bodies / Q000378:metabolism*; D011487:Protein Conformation; D017510:Protein Folding; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism*
chemical_list: D002021:Buffers; D011994:Recombinant Proteins; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; D006639:Histidine