A model of the molten globule state from molecular dynamics simulations.

Abstract

It is generally accepted that a protein's primary sequence determines its three-dmenslonal structure. It has proved difficult, however, to obtain detailed structural information about the actual protein folding process and intermediate states. We present the results of molular dynamics simulations of the unfoldin of reduced bovine pancreatic trypsin inhibitor. The resulting partially "denatured" state was compact but expanded relative to the native state (11-25%); the expansion was not caused by an influx of water molecules. The structures were mobile, with overal secondary structure contents comparable to those of the native protein.The protein experienced relatively local unfolding, with the largest changes in the structure occurring in the loop regions.A hydrophobic core was maintained althongh ping of the side chains was compromised. The properties displayed in the simulation are consistent with unfolding to a molten globule state. Our smulations provide an in-depth view of this state and details of water-protein interactions that cannot yet be obtained experimentally.A protein must assume a stable and precisely ordered conformation to perform its biological function properly. Although much is known about these conformations and their synthesis, little is known about the detailed structures and dynamic transitions that occur during protein folding. Determination of the structural characteristics of partially folded intermediate states are crucial for understanding the mechanism of folding. Unfortunately, the cooperative nature of folding results in only minute amounts of these intermediates at equilibrium. So, instead, kinetic means have been used to characterize transiently populated intermediates (1-3). However, it has been shown that a few proteins adopt stable, partially folded equilibrium intermediates under unfolding conditions (4). There is some evidence that these intermediates, termed "molten globules," also occur along the kinetic pathway (ref. 4 and references therein). However, the molecular details of even the equilibrium folding intermediates remain difficult to characterize experimentally.Here we present the results of molecular dynamics (MD) simulations of protein unfolding and the resulting partially unfolded structures. The approach of studying unfolding instead of folding has computational advantages, because simulations proceed from a well-defined starting structure. In principle, MD is capable of providing detailed information about unfolding and the resulting unfolded state that cannot be obtained experimentally. This method has yielded realistic representations ofa native protein in water (5, 6) and has been useful in understanding the nature of less-structured peptides (7-9) and the unfolding-folding transitions that they undergo (7,10,11 (14). All atoms were explicitly present during the simulations, and the potential energy function and associated parameters are based on those described earlier (15, 16), except for the parameters for chloride (R* = 2.35 A; e* = 0.10 kcal/mol...

Dynameomics: Large‐scale assessment of native protein flexibility

“…There is a nearly step-by-step correlation between high flexibility and the order of unfolding. These data suggest that native-state dynamics are closely related to unfolding and folding dynamics, in agreement with our findings in the first simulations of protein unfolding (Daggett and Levitt 1992). Later, Hespenheide et al (2002) explored the relationship between flexibility and unfolding pathways in simulations of 10 monomeric proteins and compared the results to hydrogen-deuterium exchange experiments (Li and Woodward 1999).…”

Structural Details, Pathways, and Energetics of Unfolding Apomyoglobin

“…Experimental characterization of such a barrier state is very difficult because of its extremely low population, but the presence of globally intact hydrogen bonds alongside locally disrupted packing interactions in the early unfolding of apomyoglobin 27 and DHFR 28 has been observed, and is consistent with the present simulation results, as well as with earlier MD simulations on BPTI 29 that used thermal unfolding. In their studies of partially unfolded conformations of barnase by molecular dynamics at various temperatures, Caflisch & Karplus 13 observed a delay in the entrance of water into the protein interior upon unfolding at 360 K and low pH, but no such delay at 600 K. In light of these results, our simulations at 300 K are of special interest.…”

Molecular dynamics simulations of hydrophobic collapse of ubiquitin

“…Instead, molecular dynamics simulations have focused on the unfolding process and characterization of predominantly early unfolding events (Daggett & Levitt, 1992 Tirado-Rives et al, 1997). These early kinetic unfolding events are expected to occur late in the folding process.…”