Transition state heterogeneity in GCN4 coiled coil folding studied by using multisite mutations and crosslinking.
We have investigated the folding behavior of dimeric and covalently crosslinked versions of the 33-residue alpha-helical GCN4-p1 coiled coil derived from the leucine zipper region of the transcriptional activator GCN4. The effects of multisite substitutions indicate that folding occurs along multiple routes with nucleation sites located throughout the protein. The similarity in activation energies of the different routes together with an analysis of intrinsic helical propensities indicate that minimal helix is present before a productive collision of the two chains. However, approximately one-third to one-half of the total helical structure is formed in the postcollision transition state ensemble. For the crosslinked, monomeric version, folding occurs along a single robust pathway. Here, the region nearest the crosslink, with the least helical propensity, is structured in the transition state whereas the region farthest from the tether, with the most propensity, is completely unstructured. Hence, the existence of transition state heterogeneity and the selection of folding routes critically depend on chain topology.
Moran
L B
LB
Department of Biochemistry and Molecular Biology, University of Chicago, 920 East 58th Street, Chicago, IL 60637, USA.
Schneider
J P
JP
Kentsis
A
A
Reddy
G A
GA
Sosnick
T R
TR
D000595:Amino Acid Sequence; D002467:Cell Nucleus / Q000737:chemistry; D002942:Circular Dichroism; D004268:DNA-Binding Proteins; D004305:Dose-Response Relationship, Drug; D005656:Fungal Proteins / Q000737:chemistry*; D019791:Guanidine / Q000494:pharmacology; D008954:Models, Biological; D008969:Molecular Sequence Data; D016296:Mutagenesis; D010452:Peptide Biosynthesis; D017510:Protein Folding; D011494:Protein Kinases / Q000737:chemistry*; D017433:Protein Structure, Secondary; D029701:Saccharomyces cerevisiae Proteins; D013816:Thermodynamics
D004268:DNA-Binding Proteins; D005656:Fungal Proteins; D029701:Saccharomyces cerevisiae Proteins; D011494:Protein Kinases; D019791:Guanidine