title: Proline can have opposite effects on fast and slow protein folding phases.
abstract: Proline isomerization is well known to cause additional slow phases during protein refolding. We address a new question: does the presence of prolines significantly affect the very fast kinetics that lead to the formation of folding intermediates? We examined both the very slow (10-100 min) and very fast (4 micro s-2.5 ms) folding kinetics of the two-domain enzyme yeast phosphoglycerate kinase by temperature-jump relaxation. Phosphoglycerate kinase contains a conserved cis-proline in position 204, in addition to several trans-prolines. Native cis-prolines have the largest effect on folding kinetics because the unfolded state favors trans isomerization, so we compared the kinetics of a P204H mutant with the wild-type as a proof of principle. The presence of Pro-204 causes an additional slow phase upon refolding from the cold denatured state, as reported in the literature. Contrary to this, the fast folding events are sped up in the presence of the cis-proline, probably by restriction of the conformational space accessible to the molecule. The wild-type and Pro204His mutant would be excellent models for off-lattice simulations probing the effects of conformational restriction on short timescales.
authors.affiliation: Institute of Biophysics and Radiation Biology, Semmelweis University, H-1088 Budapest, Hungary.
authors.forename: Szabolcs
authors.initials: S
authors.lastname: Osváth
authors.forename: Martin
authors.initials: M
authors.lastname: Gruebele
mesh_terms: D004789:Enzyme Activation; D004795:Enzyme Stability; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D010735:Phosphoglycerate Kinase / Q000737:chemistry*; D011392:Proline / Q000737:chemistry*; D011487:Protein Conformation; D011489:Protein Denaturation; D017510:Protein Folding; D011994:Recombinant Proteins / Q000737:chemistry; D013329:Structure-Activity Relationship; D013696:Temperature; D015003:Yeasts / Q000737:chemistry
chemical_list: D011994:Recombinant Proteins; D011392:Proline; D010735:Phosphoglycerate Kinase