title: Using deeply trapped intermediates to map the cytochrome c folding landscape.
abstract: Replacement of iron with cobalt(III) selectively introduces a deep trap in the folding-energy landscape of the heme protein cytochrome c. Remarkably, neither the protein structure nor the folding thermodynamics is perturbed by this metal-ion substitution, as shown by data from spectroscopic and x-ray diffraction experiments. Through kinetics measurements, we have found parallel folding pathways involving several different misligated Co(III) species, and, as these folding intermediates persist for several hours under certain conditions, we have been able to elucidate fully their spectroscopic properties. The results, along with an analysis of the fluorescence energy-transfer kinetics during refolding, show that rapidly equilibrating populations of compact and extended polypeptide conformations are present until all molecules have reached the native structure. These measurements provide direct evidence that collapsed denatured structures are not substantially more stable than extended conformations of cytochrome c.
authors.affiliation: Beckman Institute, California Institute of Technology, Pasadena, CA 91125, USA.
authors.forename: F Akif
authors.initials: FA
authors.lastname: Tezcan
authors.forename: William M
authors.initials: WM
authors.lastname: Findley
authors.forename: Brian R
authors.initials: BR
authors.lastname: Crane
authors.forename: Scott A
authors.initials: SA
authors.lastname: Ross
authors.forename: Julia G
authors.initials: JG
authors.lastname: Lyubovitsky
authors.forename: Harry B
authors.initials: HB
authors.lastname: Gray
authors.forename: Jay R
authors.initials: JR
authors.lastname: Winkler
mesh_terms: D000818:Animals; D002942:Circular Dichroism; D003035:Cobalt / Q000737:chemistry; D003574:Cytochrome c Group / Q000737:chemistry*; D005453:Fluorescence; D007700:Kinetics; D008958:Models, Molecular; D011487:Protein Conformation; D017510:Protein Folding; D013056:Spectrophotometry, Ultraviolet
chemical_list: D003574:Cytochrome c Group; D003035:Cobalt