Sub-microsecond protein folding.
We have investigated the structure, equilibria, and folding kinetics of an engineered 35-residue subdomain of the chicken villin headpiece, an ultrafast-folding protein. Substitution of two buried lysine residues by norleucine residues stabilizes the protein by 1 kcal/mol and increases the folding rate sixfold, as measured by nanosecond laser T-jump. The folding rate at 300 K is (0.7 micros)(-1) with little or no temperature dependence, making this protein the first sub-microsecond folder, with a rate only twofold slower than the theoretically predicted speed limit. Using the 70 ns process to obtain the effective diffusion coefficient, the free energy barrier height is estimated from Kramers theory to be less than approximately 1 kcal/mol. X-ray crystallographic determination at 1A resolution shows no significant change in structure compared to the single-norleucine-substituted molecule and suggests that the increased stability is electrostatic in origin. The ultrafast folding rate, very accurate X-ray structure, and small size make this engineered villin subdomain an ideal system for simulation by atomistic molecular dynamics with explicit solvent.
Kubelka
Jan
J
Laboratory of Chemical Physics, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD 20892-0520, USA.
Chiu
Thang K
TK
Davies
David R
DR
Eaton
William A
WA
Hofrichter
James
J
D019943:Amino Acid Substitution; D000818:Animals; D002645:Chickens; D018360:Crystallography, X-Ray; D007700:Kinetics; D008239:Lysine / Q000737:chemistry*; D008840:Microfilament Proteins / Q000737:chemistry*; D008958:Models, Molecular; D017510:Protein Folding; D017434:Protein Structure, Tertiary; D013816:Thermodynamics
D008840:Microfilament Proteins; C026615:villin; D008239:Lysine