Kinetics and thermodynamics of folding of a de Novo designed four-helix bundle protein

A simple continuum model of a lie novo designed 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 novo designed 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 Degrade 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-theta and 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-f the 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 via a kinetic partitioning mechanism. A fraction of molecules, Phi, reaches the native state by a topology inducing nucleation collapse mechanism, while the remainder (1 - Phi) follows a complex three-stage multipathway process. We suggest, in accord with our earlier studies, that Phi is essentially determined by the intrinsic temperature scales T-theta and T-f. Our studies indicate that better design of proteins can be achieved by making T-theta as close to T-f as possible. Experimental implications for de novo design of proteins are briefly discussed. (C) 1996 Academic Press Limited