Abstract
FSD-1, a designed small ultrafast folder with a ββα fold, has been actively studied in the last few years as a model system for studying protein folding mechanisms and for testing of the accuracy of computational models. The suitability of this protein to describe the folding of naturally occurring α/β proteins has recently been challenged based on the observation that the melting transition is very broad, with ill-resolved baselines. Using molecular dynamics simulations with the AMBER protein force field (ff96) coupled with the implicit solvent model (IGB = 5), we shed new light into the nature of this transition and resolve the experimental controversies. We show that the melting transition corresponds to the melting of the protein as a whole, and not solely to the helix-coil transition. The breadth of the folding transition arises from the spread in the melting temperatures (from ∼325 K to ∼302 K) of the individual transitions: formation of the hydrophobic core, β-hairpin and tertiary fold, with the helix formed earlier. Our simulations initiated from an extended chain accurately predict the native structure, provide a reasonable estimate of the transition barrier height, and explicitly demonstrate the existence of multiple pathways and multiple transition states for folding. Our exhaustive sampling enables us to assess the quality of the Amber ff96/igb5 combination and reveals that while this force field can predict the correct native fold, it nonetheless overstabilizes the α-helix portion of the protein (Tm = ∼387K) as well as the denatured structures.
Highlights
Small ‘‘ultrafast’’ folders, both naturally occurring and designed, have received considerable attention in the last few years
Small ‘‘ultrafast’’ folders, proteins that fold in microseconds, have received considerable attention, because these proteins serve as model systems for the folding of larger proteins, and permit a testing of the accuracy of computational models as well as an assessment of protein folding theories
The convergence of the replica exchange molecular dynamics (REMD) simulations was rigorously verified by a block analysis: the total sampling time of 1.25 ms for the replica at 280 K were divided into five blocks, the population of the folded structure (,3.0 Ato the NMR structure) were calculated for each block, and a good convergence was found during the 500 ns of the simulations (Text S1)
Summary
Small ‘‘ultrafast’’ folders (proteins that fold on the order of microseconds), both naturally occurring and designed, have received considerable attention in the last few years These proteins have the singular advantage of being computationally tractable, bridging the gap between experimental and in silico studies. The folding time of FSD-1 has not been reported, but the folding kinetics of a close analog, FSD-1ss (involving substitution of two non-natural aromatic residues at positions 6 and 26) have been monitored using laserinduced temperature-jump spectroscopy (Table 1) [2] This modified protein displayed two folding phases (t1,150 ns and t2,4.5 ms) at 322 K, placing FSD-1ss at the top range of known ultrafast folders.
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