Dataset Information

Probing the free energy landscapes of ALS disease mutants of SOD1 by NMR spectroscopy.

ABSTRACT: Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease that, in some cases, has been linked with mutations to the antioxidant metalloenzyme superoxide dismutase (SOD1). Although the mature form of this enzyme is highly stable and resistant to aggregation, the most immature form, lacking metal and a stabilizing intrasubunit disulfide bond, apoSOD12SH, is dynamic and hypothesized to be a major cause of toxicity in vivo. Previous solution NMR studies of wild-type apoSOD12SH have shown that the ground state interconverts with a series of sparsely populated and transiently formed conformers, some of which have aberrant nonnative structures. Here, we study seven disease mutants of apoSOD12SH and characterize their free energy landscapes as a first step in understanding the initial stages of disease progression and, more generally, to evaluate the plasticity of low-lying protein conformational states. The mutations lead to little change in the structures and dynamics of the ground states of the mutant proteins. By contrast, the numbers of low-lying excited states that are accessible to each of the disease mutants can vary significantly, with additional conformers accessed in some cases. Our study suggests that the diversity of these structures can provide alternate interaction motifs for different mutants, establishing additional pathways for new and often aberrant intra- and intermolecular contacts. Further, it emphasizes the potential importance of conformationally excited states in directing both folding and misfolding processes.

SUBMITTER: Sekhar A

PROVIDER: S-EPMC5111666 | biostudies-literature | 2016 Nov

REPOSITORIES: biostudies-literature

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Probing the free energy landscapes of ALS disease mutants of SOD1 by NMR spectroscopy.

Sekhar Ashok A Rumfeldt Jessica A O JAO Broom Helen R HR Doyle Colleen M CM Sobering Ryan E RE Meiering Elizabeth M EM Kay Lewis E LE

Proceedings of the National Academy of Sciences of the United States of America 20161024 45

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease that, in some cases, has been linked with mutations to the antioxidant metalloenzyme superoxide dismutase (SOD1). Although the mature form of this enzyme is highly stable and resistant to aggregation, the most immature form, lacking metal and a stabilizing intrasubunit disulfide bond, apoSOD1<sup>2SH</sup>, is dynamic and hypothesized to be a major cause of toxicity in vivo. Previous solution NMR studies of wild-type apoSOD1<sup> ...[more]

PMID: 27791136

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Project description:Membrane reorientation of oncogenic RAS proteins is emerging as an important modulator of their functions. Previous studies have shown that the most common orientations include those with either the three C-terminal α-helices (OS1) or N-terminal β-strands (OS2) of the catalytic domain facing the membrane. OS1 and OS2 differ by the degree to which the effector-interacting surface is occluded by the membrane. However, the relative stability of these states and the rates of transition between them remained undetermined. How mutations might modulate preferences for specific orientation states is also far from clear. The current work attempted to address these questions through a comprehensive analysis of two 20 μs-long atomistic molecular dynamics simulations. The simulations were conducted on the oncogenic G12D and Q61H KRAS mutants bound to an anionic lipid bilayer. G12D and Q61H are among the most prevalent cancer-causing mutations at the P-loop and switch 2 regions of KRAS, respectively. We found that both mutants fluctuate in a similar manner between OS1 and OS2 via an intermediate orientation OS0, and both favor the signaling competent OS1 and OS0 over the occluded OS2. However, they differ in the details, such as in the extent to which they sample OS1. Analysis of the orientation free-energy landscapes estimated from the simulations indicate that OS1 and OS2 are the most stable states. However, the overall free energy surface is rugged, indicating a large diversity of conformations including at least two substates in each orientation state that differ in stability only by about 0.5-1.0 kcal/mol. Reversible transitions between OS1 and OS2 occur via two well-defined pathways that traverse OS0. In the minimum energy path, helix 4 remains close to the membrane as the angle of the catalytic domain from the membrane plane changes, resulting in a barrier of ∼1 kcal/mol for OS1/OS2 interconversions. Estimation of the rates of the various transitions based on survival probabilities yielded two rate constants in the order of 107 and 106 s-1, which we attribute to intrinsic protein conformational dynamics and transient protein-lipid interactions, respectively. The faster process dominates every transition, confirming a previous suggestion that RAS membrane reorientation is driven by conformational fluctuations rather than protein-lipid interactions.

Dataset Information

Probing the free energy landscapes of ALS disease mutants of SOD1 by NMR spectroscopy.

Publications

Probing the free energy landscapes of ALS disease mutants of SOD1 by NMR spectroscopy.

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