Kinetics unfolding

We carried out unfolding MD simulations of goat a-lactalbumin at 498 K. The protein structure was represented in the segmental Q-coordinate, and cluster analyses and multiple-trajectory alignments were carried out to obtain the transition-state structure solely from the MD simulation. The structure obtained by this approach was very close to that obtained experimentally, and hence the results of the kinetic unfolding experiments were well reproduced by the simulations. 

At the macroscopic scale, the full competition of reactant diffusion, electron and proton migration, and charge transfer kinetics unfolds. The water balance further complicates this interplay. Moreover, performance is subject to operation conditions and complex boundary conditions at interfaces to membrane and gas diffusion layer. A vast list of structural characteristics steers this interplay, including thickness, composition, pore size distributions, and wetting properties of pores. 

Wilson, D.J., Rafferty, S.P., Konermann, L. (2005) Kinetic Unfolding Mechanism of the Inducible Nitric Oxide Synthase Oxygenase Domain Determined by Time-resolved Electrospray Mass Spectrometry. Biochemistry 44 2276-2283. 

Given that a sequence folds to a known native stmcture, what are the mechanisms in the transition from the unfolded confonnation to the folded state This is a kinetics problem, the solution of which requires elucidation of the pathways and transition states in the folding process. 

For these sequences the value of Gj, is less than a certain small value g. For such sequences the folding occurs directly from the ensemble of unfolded states to the NBA. The free energy surface is dominated by the NBA (or a funnel) and the volume associated with NBA is very large. The partition factor <6 is near unify so that these sequences reach the native state by two-state kinetics. The amplitudes in (C2.5.7) are nearly zero. There are no intennediates in the pathways from the denatured state to the native state. Fast folders reach the native state by a nucleation-collapse mechanism which means that once a certain number of contacts (folding nuclei) are fonned then the native state is reached very rapidly [25, 26]. The time scale for reaching the native state for fast folders (which are nonnally associated with those sequences for which topological fmstration is minimal) is found to be... 

Measuring Protein Sta.bihty, Protein stabihty is usually measured quantitatively as the difference in free energy between the folded and unfolded states of the protein. These states are most commonly measured using spectroscopic techniques, such as circular dichroic spectroscopy, fluorescence (generally tryptophan fluorescence) spectroscopy, nmr spectroscopy, and absorbance spectroscopy (10). For most monomeric proteins, the two-state model of protein folding can be invoked. This model states that under equihbrium conditions, the vast majority of the protein molecules in a solution exist in either the folded (native) or unfolded (denatured) state. Any kinetic intermediates that might exist on the pathway between folded and unfolded states do not accumulate to any significant extent under equihbrium conditions (39). In other words, under any set of solution conditions, at equihbrium the entire population of protein molecules can be accounted for by the mole fraction of denatured protein, and the mole fraction of native protein,, ie. 

Solving the master equation for the minimally frustrated random energy model showed that the kinetics depend on the connectivity [23]. Eor the globally connected model it was found that the resulting kinetics vary as a function of the energy gap between the folded and unfolded states and the roughness of the energy landscape. The model... 

Raschke, T. M., and Marqnsee, S., 1997. The kinetic folding intermediate of ribonnclease H resembles the acid molten globule and partially unfolded molecules detected under native conditions. Nature Structural Biology 4 298-304. 

K. Benedek, S. Dong and B. E. Kaiger, Kinetics of unfolding of proteins on hydrophobic surfaces in reversed-phase liquid chiomatography , /. Chromatogr. 317 227-243 (1984). 

We will limit ourselves to reviewing recent SAXS and SANS studies of putatively fully unfolded states formed at equilibrium. We direct readers interested in partially folded states (kinetic and equilibrium molten globules and their brethren) to a number of excellent recent articles and reviews (Kataoka and Goto, 1996 Kataoka et al., 1997 Uversky etal., 1998 Pollack et al., 1999 Doniach, 2001). Similarly, we will not discuss in detail the technical aspects of scattering studies or the precise interpretation of scattering profiles, but instead direct the reader to the appropriate resources (Glatter and Kratky, 1982 Doniach et al., 1995 Kataoka and Goto, 1996 Doniach, 2001). 

Polypeptide chains exist in an equilibrium between different conformations as a function of environment (solvent, other solutes, pH) and thermodynamic (temperature, pressure) conditions. If a polypeptide adopts a structurally ordered, stable conformation, one speaks of an equilibrium between a folded state, represented by the structured, densely populated conformer, and an unfolded state, represented by diverse, sparsely populated conformers. Although this equilibrium exists for polypeptide chains of any size, its thermodynamics and kinetics are typically different for oligopeptides and proteins. This can be broadly explained with reference to the different dimensionalities of the free-energy hypersurfaces of these two types of molecules. 

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