Functional Domain Motions in Proteins on the ∼1–100 ns Timescale: Comparison of Neutron Spin-Echo Spectroscopy of Phosphoglycerate Kinase with Molecular-Dynamics Simulation

University of Tennessee/Oak Ridge National Laboratory Center for Molecular Biophysics, Oak Ridge National Laboratory, Oak Ridge, Tennessee, USA.
Biophysical Journal (Impact Factor: 3.97). 03/2012; 102(5):1108-17. DOI: 10.1016/j.bpj.2012.01.002
Source: PubMed


Protein function often requires large-scale domain motion. An exciting new development in the experimental characterization of domain motions in proteins is the application of neutron spin-echo spectroscopy (NSE). NSE directly probes coherent (i.e., pair correlated) scattering on the ~1-100 ns timescale. Here, we report on all-atom molecular-dynamics (MD) simulation of a protein, phosphoglycerate kinase, from which we calculate small-angle neutron scattering (SANS) and NSE scattering properties. The simulation-derived and experimental-solution SANS results are in excellent agreement. The contributions of translational and rotational whole-molecule diffusion to the simulation-derived NSE and potential problems in their estimation are examined. Principal component analysis identifies types of domain motion that dominate the internal motion's contribution to the NSE signal, with the largest being classic hinge bending. The associated free-energy profiles are quasiharmonic and the frictional properties correspond to highly overdamped motion. The amplitudes of the motions derived by MD are smaller than those derived from the experimental analysis, and possible reasons for this difference are discussed. The MD results confirm that a significant component of the NSE arises from internal dynamics. They also demonstrate that the combination of NSE with MD is potentially useful for determining the forms, potentials of mean force, and time dependence of functional domain motions in proteins.

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Available from: Gerald Kneller, Oct 04, 2015
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    • "In the absence of substrates PGK remains in a wide open conformation with ligand binding sites fully exposed to the solvent . Recent SANS and NSE studies have shown that the enzyme is subject to Brownian forces that will drive domain closure on a timescale of 50 ns, a rate that is in accord with kinetic turnover rates [52] [53]. When domain closure occurs, electrostatic forces then come into play. "
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    ABSTRACT: Domain motions are essential to many catalytic mechanisms in enzymes but they are often difficult to study. X-ray crystal structures can provide molecular details of snapshots of catalysis but many states important in the cycle remain inaccessible using this technique. Phosphoglycerate kinase (PGK) undergoes large domain movements in order to catalyse the production of ATP. PGK is the enzyme responsible for the first ATP generating step of glycolysis and has been implicated in oncogenesis and the in vivo activation of l-nucleoside pro-drugs effective against retroviruses. Its mechanism requires considerable hinge bending to bring the substrates into proximity in order for phosphoryl transfer to occur. The enzyme has been the subject of intense study for decades but new crystal structures, methods in solution scattering and modelling techniques are throwing light on the dynamics of catalysis of this archetypal kinase. Here, I argue that Brownian forces acting on the protein are the dominant factor in the catalytic cycle and that the enzyme has evolved measures to harness this force for efficient catalysis.
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