Fast folding of a helical protein initiated by the collision of unstructured chains

Abstract

To examine whether helix formation necessarily precedes chain collision, we have measured the folding of a fully helical coiled coil that has been specially engineered to have negligible intrinsic helical propensity but high overall stability. The folding rate approaches the diffusion-limited value and is much faster than possible if folding is contingent on precollision helix formation. Therefore, the collision of two unstructured chains is the initial step of the dominant kinetic pathway, whereas helicity exerts its influence only at a later step. Folding from an unstructured encounter complex may be efficient and robust, which has implications for any biological process that couples folding to binding.ne of the most debated issues in protein folding concerns the earliest folding events leading up to the transition state (1-9). For helical proteins, the earliest productive folding steps often are postulated to involve the collision of two preformed, but not necessarily stable, helical elements, rather than collision of unstructured chains. This diffusion-collision model (D-C model) (10-12) is supported by the observation that helix formation is faster than overall folding rates (13-15). This broadly accepted view also is supported by the presence of helix in the folding transition state and an increase in k f with an increase in helical propensity (2, 4, 16-21).However, this correlation can support an opposing model in which unstructured chains first collide, and the enhanced helicity increases the success frequency (or transmission coefficient) of each encounter (Fig. 1). In general, the highly cooperative (two-state) folding behavior of most small proteins precludes identifying the order of events leading up to the kinetic barrier. As a result, the demonstration of helical structure in the transition state cannot by itself resolve whether helix formation or chain collision occurs first.This obstacle can be overcome by studying a system with minimal helical propensity and composed of more than one chain, so that the rate of collision can be varied (22). These properties enable comparisons between observed folding rates and the maximum rate consistent with a model where precollision helix formation is required. Our investigation uses this strategy in conjunction with a dimeric coiled coil protein specially engineered to have negligible intrinsic helicity but high stability. This protein folds at nearly the diffusion limit and certainly at a much faster rate than would be possible if the helix must form before collision. Thus, an unstructured encounter complex can successfully initiate rapid folding, with helix formation occurring at a later step. The collision-first route sets a high basal level for the folding rate of any protein.

The loop hypothesis: contribution of early formed specific non-local interactions to the determination of protein folding pathways

“…Kato et al (2010) studied the folding of mutants of S. nuclease, where NLIs were perturbed and concluded that the native NLIs established at the early stage of the folding process facilitate further secondary structure formation. Meisner and Sosnick (2004) studied the kinetics of folding of a two-chain engineered protein in which helicity and collision rate can be varied and concluded that an unstructured encounter complex can successfully initiate rapid folding, with helix formation occurring at a later stage. The collision-first route enables a high basal folding rate of any protein.…”

Molecular basis of coiled-coil formation

“…Along these lines, we have been interested particularly in coiled-coil ''trigger sequences,'' which encode stable monomeric ␣-helices that are indispensable for coiled-coil formation (14)(15)(16)(17)(18). Although there are a few examples of synthetic peptides that fold either into heterodimers (19) or at conditions of extremes of pH (20) without an apparent trigger sequence, the ''trigger site'' concept is generally accepted because these short autonomous helical folding units are structurally and functionally conserved in a large number of native coiled-coil proteins (reviewed in refs. [21][22][23][24].…”

Improved Stability of the Jun-Fos Activator Protein-1 Coiled Coil Motif

“…Indeed, a wealth of reports have been published on the folding of the yeast transcription factor GCN4-p1 (12,14,(17)(18)(19)(20)(21) with many reporting the protein to fold in a two-state mechanism. However, some have speculated about the existence of intermediates (22)(23)(24).…”