Solvent-tuning the collapse and helix formation time scales of lambda(6-85)*.
The lambda(6-85)(*) pseudo-wild type of lambda repressor fragment is a fast two-state folder (k(f) approximately 35 microsec(-1) at 58 degrees C). Previously, highly stable lambda(6-85)(*) mutants with k(f) > 30 microsec(-1) have been engineered to fold nearly or fully downhill. Stabilization of the native state by solvent tuning might also tune lambda(6-85)(*) away from two-state folding. We test this prediction by examining the folding thermodynamics and kinetics of lambda(6-85)(*) in a stabilizing solvent, 45% by weight aqueous ethylene glycol at -28 degrees C. Detection of kinetics by circular dichroism at 222 nm (sensitive to helix content) and small angle X-ray scattering (measuring the radius of gyration) shows that refolding from guanidine hydrochloride denatured conditions exhibits very different time scales for collapse and secondary structure formation: the two processes become decoupled. Collapse remains a low-barrier activated process, while the fastest of several secondary structure formation time scales approaches the downhill folding limit. Two-state folding of lambda(6-85)(*) is not a robust process.
Dumont
Charles
C
Department of Physics, University of Illinois at Urbana-Champaign, Urbana, Illinois 61801, USA.
Matsumura
Yoshitaka
Y
Kim
Seung Joong
SJ
Li
Jinsong
J
Kondrashkina
Elena
E
Kihara
Hiroshi
H
Gruebele
Martin
M
D002942:Circular Dichroism / Q000379:methods; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000187:drug effects*; D008958:Models, Molecular; D050505:Mutant Proteins / Q000737:chemistry; D017510:Protein Folding; D017433:Protein Structure, Secondary / Q000187:drug effects*; D012097:Repressor Proteins / Q000737:chemistry* / Q000187:drug effects*; D012997:Solvents / Q000494:pharmacology*; D013816:Thermodynamics; D014764:Viral Proteins / Q000737:chemistry* / Q000187:drug effects*; D054334:Viral Regulatory and Accessory Proteins; D014961:X-Ray Diffraction / Q000379:methods
D004268:DNA-Binding Proteins; D050505:Mutant Proteins; D012097:Repressor Proteins; D012997:Solvents; D014764:Viral Proteins; D054334:Viral Regulatory and Accessory Proteins; C016649:phage repressor proteins