title: raf RBD and ubiquitin proteins share similar folds, folding rates and mechanisms despite having unrelated amino acid sequences.
abstract: Recent experimental and theoretical studies in protein folding suggest that the rates and underlying mechanisms by which proteins attain the native state are largely determined by the topological complexity of a specific fold rather than by the fine details of the amino acid sequences. However, such arguments are based upon the examination of a limited number of protein folds. To test this view, we sought to investigate whether proteins belonging to the ubiquitin superfamily display similar folding behavior. To do so, we compared the folding-unfolding transitions of mammalian ubiquitin (mUbi) with those of its close yeast homologue (yUbi), and to those of the structurally related Ras binding domain (RBD) of the serine/threonine kinase raf that displays no apparent sequence homology with the ubiquitin family members. As demonstrated for mUbi [Krantz, B. A., and Sosnick, T. R. (2000) Biochemistry 39, 11696-11701], we show that a two-state transition model with no burst phase intermediate can describe folding of both yUbi and raf RBD. We further demonstrate that (1) all three proteins refold at rates that are within 1 order of magnitude (1800, 1100, and 370 s(-1) for mUbi, raf RBD, and yUbi, respectively), (2) both mUbi and raf RBD display similar refolding heterogeneity, and (3) the folding free energy barriers of both mUbi and raf RBD display a similar temperature dependence and sensitivity to a stabilizing agent or to mutations of a structurally equivalent central core residue. These findings are consistent with the view that rates and mechanisms for protein folding depend mostly on the complexity of the native structure topology rather than on the fine details of the amino acid sequence.
authors.affiliation: Département de biochimie, Université de Montréal, CP 6128, Station Centre-Ville, Montréal, Québec H3C 3J7, Canada.
authors.forename: Alexis
authors.initials: A
authors.lastname: Vallée-Bélisle
authors.forename: Jean-François
authors.initials: JF
authors.lastname: Turcotte
authors.forename: Stephen W
authors.initials: SW
authors.lastname: Michnick
mesh_terms: D000595:Amino Acid Sequence; D000596:Amino Acids / Q000737:chemistry; D004305:Dose-Response Relationship, Drug; D007700:Kinetics; D008956:Models, Chemical; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D011487:Protein Conformation; D017510:Protein Folding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D019908:Proto-Oncogene Proteins c-raf / Q000737:chemistry*; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D013696:Temperature; D013816:Thermodynamics; D025801:Ubiquitin / Q000737:chemistry
chemical_list: D000596:Amino Acids; D025801:Ubiquitin; D019908:Proto-Oncogene Proteins c-raf