Characterizing protein folding transition States using Psi-analysis.
We discuss the implementation of Psi-analysis for the structural characterization of protein folding transition states. In Psi-analysis, engineered bi-histidine metal ion binding sites are introduced at surface positions to stabilize secondary and tertiary structures. The addition of metal ions stabilizes the interaction between the two known histidines in a continuous fashion. Measuring the ratio of transition state stabilization to that of the native state provides information about the presence of the metal binding site in the transition state. Psi-Analysis uses noninvasive surface mutations and does not require specialized equipment, so it can be readily applied to characterize the folding of many proteins. As a result, this method can provide a wealth of high-resolution quantitative data for comparison with theoretical folding simulations. Additionally, investigations of other biological processes also may utilize metal binding sites and Psi-analysis to detect conformational events during catalysis, assembly, and function.
Pandit
Adarsh D
AD
Department of Biochemistry and Molecular Biology, University of Chicago, IL, USA.
Krantz
Bryan A
BA
Dothager
Robin S
RS
Sosnick
Tobin R
TR
D001665:Binding Sites; D006639:Histidine / Q000737:chemistry; D007700:Kinetics; D008670:Metals / Q000737:chemistry; D008958:Models, Molecular; D011487:Protein Conformation; D015202:Protein Engineering / Q000379:methods*; D017510:Protein Folding; D011506:Proteins / Q000737:chemistry*
D008670:Metals; D011506:Proteins; D006639:Histidine