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Separating instability from aggregation propensity in ?S-crystallin variants.

ABSTRACT: Molecular dynamics (MD) simulations, circular dichroism (CD), and dynamic light scattering (DLS) measurements were used to investigate the aggregation propensity of the eye-lens protein ?S-crystallin. The wild-type protein was investigated along with the cataract-related G18V variant and the symmetry-related G106V variant. The MD simulations suggest that local sequence differences result in dramatic differences in dynamics and hydration between these two apparently similar point mutations. This finding is supported by the experimental measurements, which show that although both variants appear to be mostly folded at room temperature, both display increased aggregation propensity. Although the disease-related G18V variant is not the most strongly destabilized, it aggregates more readily than either the wild-type or the G106V variant. These results indicate that ?S-crystallin provides an excellent model system for investigating the role of dynamics and hydration in aggregation by locally unfolded proteins.

SUBMITTER: Brubaker WD

PROVIDER: S-EPMC3021659 | biostudies-literature | 2011 Jan

REPOSITORIES: biostudies-literature

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Separating instability from aggregation propensity in γS-crystallin variants.

Brubaker William D WD Freites J Alfredo JA Golchert Kory J KJ Shapiro Rebecca A RA Morikis Vasilios V Tobias Douglas J DJ Martin Rachel W RW

Biophysical journal 20110101 2

Molecular dynamics (MD) simulations, circular dichroism (CD), and dynamic light scattering (DLS) measurements were used to investigate the aggregation propensity of the eye-lens protein γS-crystallin. The wild-type protein was investigated along with the cataract-related G18V variant and the symmetry-related G106V variant. The MD simulations suggest that local sequence differences result in dramatic differences in dynamics and hydration between these two apparently similar point mutations. This ...[more]

PMID: 21244846

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Project description:BackgroundThe eye lens crystallins are highly soluble proteins that are required to last the lifespan of an organism due to low protein turnover in the lens. Crystallin aggregation leads to formation of light-scattering aggregates known as cataract. The G18V mutation of human ?S-crystallin (?S-G18V), which is associated with childhood-onset cataract, causes structural changes throughout the N-terminal domain and increases aggregation propensity. The holdase chaperone protein ?B-crystallin does not interact with wild-type ?S-crystallin, but does bind its G18V variant. The specific molecular determinants of ?B-crystallin binding to client proteins is incompletely charcterized. Here, a new variant of ?S, ?S-G18A, was created to test the limits of ?B-crystallin selectivity.MethodsMolecular dynamics simulations were used to investigate the structure and dynamics of ?S-G18A. The overall fold of ?S-G18A was assessed by circular dichroism (CD) spectroscopy and intrinsic tryptophan fluorescence. Its thermal unfolding temperature and aggregation propensity were characterized by CD and DLS, respectively. Solution-state NMR was used to characterize interactions between ?B-crystallin and ?S-G18A.Results?S-G18A exhibits minimal structural changes, but has compromised thermal stability relative to ?S-WT. The placement of alanine, rather than valine, at this highly conserved glycine position produces minor changes in hydrophobic surface exposure. However, human ?B-crystallin does not bind the G18A variant, in contrast to previous observations for ?S-G18V, which aggregates at physiological temperature.Conclusions?B-crystallin is capable of distinguishing between aggregation-prone and function-preserving variants, and recognizing the transient unfolding or minor conformers that lead to aggregation in the disease-related variant.General significanceHuman ?B-crystallin distinguishes between highly similar variants of a structural crystallin, binding the cataract-related ?S-G18V variant, but not the function-preserving ?S-G18A variant, which is monomeric at physiological temperature.

Dataset Information

Separating instability from aggregation propensity in ?S-crystallin variants.

Publications

Separating instability from aggregation propensity in γS-crystallin variants.

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