Consequences of localized frustration for the folding mechanism of the IM7 protein

Abstract

In the laboratory, IM7 has been found to have an unusual folding mechanism in which an ''on-pathway'' intermediate with nonnative interactions is formed. We show that this intermediate is a consequence of an unusual cluster of highly frustrated interactions in the native structure. This cluster is involved in the binding of IM7 to its target, Colicin E7. Redesign of residues in this cluster to eliminate frustration is predicted by simulations to lead to faster folding without the population of an intermediate ensemble.computational design ͉ folding landscape ͉ nonnative intermediate F or proteins to fold to an organized native ensemble, they have evolved to form a highly favorable, stabilizing set of native interactions (1). Proteins, it seems, have a funneled energy landscape toward the native state (1-3). However, real protein landscapes may not be perfectly funneled but probably display some landscape ruggedness because of the possibility of forming favorable but nonnative interactions during the folding process. Energy landscape theory shows that, if the landscape were too rugged, numerous long-lived intermediate states that energetically compete with the native state would become populated (4). The rarity of the experimental finding of such intermediates having significant nonnative structure strongly argues that proteins largely conform to the ''principle of minimal frustration'' (2).When populated kinetic folding intermediates have been observed, it has generally turned out to be the case that these metastable thermodynamic states are not caused by energetic frustration but by a nonuniform compensation of the entropy and energy changes upon forming native contacts. The existence of such productive, on route, folding intermediates can be said to be topology-driven and can often be predicted from the native structure alone by using a perfectly funneled landscape (5-8).IM7 is an 86-residue-long protein that is known to fold through an on-pathway intermediate state. An intermediate populated in the folding of IM7 has been shown to possess a nonnative packing of three of the four helices around a specific hydrophobic core (9). In this article, we explore the relationship between nonnative interactions, the presence of the stable folding intermediate, and the residual frustration found in the native structure. Our analysis suggests that this seeming exception to the pattern expected for minimally frustrated proteins is, in fact, consistent with the ideas of energy landscape theory touching on the behavior of frustrated, random heteropolymers.We first demonstrate that the intermediate of IM7 is not a direct consequence of the topology of IM7 by simulating its folding with a sequence-independent energy function that yields a perfectly funneled landscape [a so-called Go Hamiltonian (10) Capaldi et al. (9), who inferred the presence of a three-helix core of helices I, II, and IV. We note that this ensemble has numerous native-like interactions but also is stabilized by some nonnative interactions.Equipped with an ...

Theoretical and experimental demonstration of the importance of specific nonnative interactions in protein folding

“…In a similar vein, local propensity for a nonnative backbone conformation was found to be a strong kinetic driving force in the folding of Fyn SH3 domain, indicating that its folding transition state contains a specific nonnative conformation (17). These experimental findings confirmed general predictions from simulations (18)(19)(20)(21)(22) and analytical theory (20,23) that nonnative interactions can accelerate folding under certain conditions and led to the concept of ''localized frustration'' as a possible basis for the nonnative interactions in the folding intermediate of the Im7 protein (24). However, despite these advances, a direct comparison between theory and experiment on the folding effects of specific nonnative interactions has not been performed.…”

Localizing frustration in native proteins and protein assemblies

“…Our statistical survey has shown that these sites do colocalize with regions involved in the formation of heterodimeric protein assemblies. An accompanying article in this issue of PNAS (27) describes in detail a particularly interesting application of these concepts highlighting the role played by local frustration in the folding mechanism of the Im7 protein.…”

On the reliability of peptide nonplanarity seen in ultra‐high resolution crystal structures

“…The standard deviation was 6.38 for trans peptide bonds with about 12% and 0.5% of residues deviating more than 108and 208, respectively, from planarity. As not all parts of a crystal structure have the same level of reliability, we examined the electron density of every peptide with x 208 from planar to assess which were reliable; then using the reliable examples, we showed that these highly nonplanar peptides are not just in active sites, but represent a mundane aspect of protein structure that simply reflects the "frustration" [16][17][18] that occurs as proteins fold into their tertiary structures. We (see Fig.…”