title: Acceleration of the Rate-Limiting Step of Thioredoxin Folding by Replacement of its Conserved cis-Proline with (4 S)-Fluoroproline.
abstract: The incorporation of the non-natural amino acids (4R)- and (4S)-fluoroproline (Flp) has been successfully used to improve protein stability, but little is known about their effect on protein folding kinetics. Here we analyzed the influence of (4R)- and (4S)-Flp on the rate-limiting trans-to-cis isomerization of the Ile75-Pro76 peptide bond in the folding of Escherichia coli thioredoxin (Trx). While (4R)-Flp at position 76 had essentially no effect on the isomerization rate in the context of the intact tertiary structure, (4S)-Flp accelerated the folding reaction ninefold. Similarly, tenfold faster trans-to-cis isomerization of Ile75-(4S)-Flp76 relative to Ile75-Pro76 was observed in the unfolded state of Trx. Our results show that the replacement of cis prolines by non-natural proline analogues can be used for modulating the folding rates of proteins with cis prolyl-peptide bonds in the native state.
authors.affiliation: Department of Biology, Institute of Molecular Biology and Biophysics, ETH Zürich, Otto-Stern-Weg 5, 8093, Zürich, Switzerland.|Max-Planck-Institute of Molecular Physiology, Department of Structural Biochemistry, Otto-Hahn-Strasse 11, 44227, Dortmund, Germany.
authors.forename: Daniel
authors.initials: D
authors.lastname: Roderer
authors.affiliation: Department of Biology, Institute of Molecular Biology and Biophysics, ETH Zürich, Otto-Stern-Weg 5, 8093, Zürich, Switzerland.
authors.forename: Rudi
authors.initials: R
authors.lastname: Glockshuber
authors.affiliation: Department of Organic Chemistry, University of Konstanz, Universitätsstrasse 10, 78464, Konstanz, Germany. marina.rubini@uni-konstanz.de.
authors.forename: Marina
authors.initials: M
authors.lastname: Rubini
mesh_terms: D004926:Escherichia coli / Q000737:chemistry; D008958:Models, Molecular; D011392:Proline / Q000031:analogs & derivatives* / Q000737:chemistry* / Q000378:metabolism; D017510:Protein Folding; D013237:Stereoisomerism; D013879:Thioredoxins / Q000737:chemistry*
chemical_list: C431803:fluoro-proline; D013879:Thioredoxins; D011392:Proline
keywords: amino acids; cis/trans isomerization; fluoroproline; folding kinetics; protein folding; thioredoxin