Kinetics and Thermodynamics of Folding of a de NovoDesigned Four-helix Bundle Protein

Abstract

A simple continuum model of a de novodesigned model of a four-helix bundle is presented. The thermodynamics and kinetics of the model are studied using Langevin simulations. We use a three-letter minimal off- lattice representation of a de novodesigned four-helix bundle protein. The native state of the model, which can be thought of as an alpha-carbon representation of the peptide chain, is a caricature of the sequence designed by Ho and Degrado and shows several characteristics found in the naturally occurring four-helix bundles. These include the structural aspects and the relative stability of the native conformation. The model four-helix bundle shows two characteristic temperatures T θand T f. The former is the temperature above which the structure resembles that of the random coil. Below the first-order folding transition temperature T fthe chain adopts the native conformation corresponding to the four-helix bundle. It is shown that in order to obtain a unique native structure a proper free energy balance between secondary and tertiary interactions is needed. The thermal denaturation starting from the unique native conformation indicates that at least a three-state analysis is required. The intermediates in the equilibrium thermal denaturation are all found to be native-like. The kinetics of refolding starting from an ensemble of denatured states shows that the acquisition of the native conformation takes place viaa kinetic partitioning mechanism. A fraction of molecules, Φ, reaches the native state by a topology inducing nucleation collapse mechanism, while the remainder (1 −Φ) follows a complex three-stage multipathway process. We suggest, in accord with our earlier studies, that Φ is essentially determined by the intrinsic temperature scales T θand T f. Our studies indicate that better design of proteins can be achieved by making T θas close to T fas possible. Experimental implications for de novodesign of proteins are briefly discussed.