To further investigate the folding mechanism of the villin subdomian, we have studied the equilibrium and kinetics using laser temperature jump of 8 additional mutants in which the amino replacement contains a simple methyl group deletion. In most cases this results in a non-natural amino acid, but the protein is made by peptide synthesis so this does not pose a problem. Instead of calculating phi values, the equilibrium curves are being fit with the Ising-like model, and the new relaxation rate is calculated on the perturbed free energy surface to compare with the measured relaxation rate. The most important mutant is one in which 2 buried lysines have been replaced by norleucines. These mutations eliminate 2 repulsive electrostatic interactions, stabilize the protein, and speed up folding to 700 nanoseconds - the current world record for folding speed. This molecule is being intensively investigated by the major molecular dynamics groups working on protein folding. In collaboration with Dr. Marco Buscaglia of the University of Milan, a former post-doc, we have confirmed the rate of 1/(700 ns) using a totally independent method that use tryptophan quenching by cysteine.
| Cellmer, Troy; Buscaglia, Marco; Henry, Eric R et al. (2011) Making connections between ultrafast protein folding kinetics and molecular dynamics simulations. Proc Natl Acad Sci U S A 108:6103-8 |
| Cellmer, Troy; Henry, Eric R; Hofrichter, James et al. (2008) Measuring internal friction of an ultrafast-folding protein. Proc Natl Acad Sci U S A 105:18320-5 |
| Kubelka, Jan; Henry, Eric R; Cellmer, Troy et al. (2008) Chemical, physical, and theoretical kinetics of an ultrafast folding protein. Proc Natl Acad Sci U S A 105:18655-62 |
