The folding trajectory of RNase H is dominated by its topology and not local stability: a protein engineering study of variants that fold via two-state and three-state mechanisms.
Proteins can sample a variety of partially folded conformations during the transition between the unfolded and native states. Some proteins never significantly populate these high-energy states and fold by an apparently two-state process. However, many proteins populate detectable, partially folded forms during the folding process. The role of such intermediates is a matter of considerable debate. A single amino acid change can convert Escherichia coli ribonuclease H from a three-state folder that populates a kinetic intermediate to one that folds in an apparent two-state fashion. We have compared the folding trajectories of the three-state RNase H and the two-state RNase H, proteins with the same native-state topology but altered regional stability, using a protein engineering approach. Our data suggest that both versions of RNase H fold through a similar trajectory with similar high-energy conformations. Mutations in the core and the periphery of the protein affect similar aspects of folding for both variants, suggesting a common trajectory with folding of the core region followed by the folding of the periphery. Our results suggest that formation of specific partially folded conformations may be a general feature of protein folding that can promote, rather than hinder, efficient folding.
Connell
Katelyn B
KB
Chemical Biology Graduate Group, Department of Chemistry, Institute for Quantitative Biosciences-Berkeley, University of California, Berkeley, CA 94720-3220, USA.
Miller
Erik J
EJ
Marqusee
Susan
S
D004795:Enzyme Stability; D004926:Escherichia coli / Q000201:enzymology*; D009154:Mutation; D011487:Protein Conformation; D015202:Protein Engineering; D017510:Protein Folding; D016914:Ribonuclease H / Q000737:chemistry* / Q000235:genetics
D016914:Ribonuclease H; C078856:ribonuclease HI