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The congenital cataract-linked G61C mutation destabilizes γD-crystallin and promotes non-native aggregation.

Zhang W, Cai HC, Li FF, Xi YB, Ma X, Yan YB - PLoS ONE (2011)

Bottom Line: The stability of γD-crystallin against heat- or GdnHCl-induced denaturation was significantly decreased by the mutation, while no influence was observed on the acid-induced unfolding.The aggregation-prone property of the mutant was not altered by the addition of reductive reagent.These results suggested that the decrease in protein stability followed by aggregation-prone property might be the major cause in the hereditary cataract induced by the G61C mutation.

View Article: PubMed Central - PubMed

Affiliation: State Key Laboratory of Biomembrane and Membrane Biotechnology, School of Life Sciences, Tsinghua University, Beijing, China.

ABSTRACT
γD-crystallin is one of the major structural proteins in human eye lens. The solubility and stability of γD-crystallin play a crucial role in maintaining the optical properties of the lens during the life span of an individual. Previous study has shown that the inherited mutation G61C results in autosomal dominant congenital cataract. In this research, we studied the effects of the G61C mutation on γD-crystallin structure, stability and aggregation via biophysical methods. CD, intrinsic and extrinsic fluorescence spectroscopy indicated that the G61C mutation did not affect the native structure of γD-crystallin. The stability of γD-crystallin against heat- or GdnHCl-induced denaturation was significantly decreased by the mutation, while no influence was observed on the acid-induced unfolding. The mutation mainly affected the transition from the native state to the intermediate but not that from the intermediate to the unfolded or aggregated states. At high temperatures, both proteins were able to form aggregates, and the aggregation of the mutant was much more serious than the wild type protein at the same temperature. At body temperature and acidic conditions, the mutant was more prone to form amyloid-like fibrils. The aggregation-prone property of the mutant was not altered by the addition of reductive reagent. These results suggested that the decrease in protein stability followed by aggregation-prone property might be the major cause in the hereditary cataract induced by the G61C mutation.

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Phase diagram analysis of the intrinsic fluorescence spectra of the WT (filled symbols and solid lines) and mutated γD-crystallin (open symbols and dashed lines) during thermal unfolding.The phase diagram is constructed by monitoring the changes of the fluorescence intensity at 365 nm (I365) as a function of that at 320 nm (I320). The temperatures of the starting point, the joint position of the adjacent lines and the end point are labeled.
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pone-0020564-g004: Phase diagram analysis of the intrinsic fluorescence spectra of the WT (filled symbols and solid lines) and mutated γD-crystallin (open symbols and dashed lines) during thermal unfolding.The phase diagram is constructed by monitoring the changes of the fluorescence intensity at 365 nm (I365) as a function of that at 320 nm (I320). The temperatures of the starting point, the joint position of the adjacent lines and the end point are labeled.

Mentions: To further investigate the sequential events occurred during γD-crystallin thermal denaturation, phase diagram was constructed to check whether there existed an unfolding intermediate. In the phase diagram, each linear part indicates a two-state process, and the joint-point of two adjacent lines indicates the appearance of a folding intermediate at this position [41]. As shown in Figure 4, the phase diagram of both the WT and mutated proteins involved two linear parts but different joint positions, confirming that the thermal unfolding of both proteins involved an intermediate: native state (N)→intermediate (I)→aggregates (A).


The congenital cataract-linked G61C mutation destabilizes γD-crystallin and promotes non-native aggregation.

Zhang W, Cai HC, Li FF, Xi YB, Ma X, Yan YB - PLoS ONE (2011)

Phase diagram analysis of the intrinsic fluorescence spectra of the WT (filled symbols and solid lines) and mutated γD-crystallin (open symbols and dashed lines) during thermal unfolding.The phase diagram is constructed by monitoring the changes of the fluorescence intensity at 365 nm (I365) as a function of that at 320 nm (I320). The temperatures of the starting point, the joint position of the adjacent lines and the end point are labeled.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3105094&req=5

pone-0020564-g004: Phase diagram analysis of the intrinsic fluorescence spectra of the WT (filled symbols and solid lines) and mutated γD-crystallin (open symbols and dashed lines) during thermal unfolding.The phase diagram is constructed by monitoring the changes of the fluorescence intensity at 365 nm (I365) as a function of that at 320 nm (I320). The temperatures of the starting point, the joint position of the adjacent lines and the end point are labeled.
Mentions: To further investigate the sequential events occurred during γD-crystallin thermal denaturation, phase diagram was constructed to check whether there existed an unfolding intermediate. In the phase diagram, each linear part indicates a two-state process, and the joint-point of two adjacent lines indicates the appearance of a folding intermediate at this position [41]. As shown in Figure 4, the phase diagram of both the WT and mutated proteins involved two linear parts but different joint positions, confirming that the thermal unfolding of both proteins involved an intermediate: native state (N)→intermediate (I)→aggregates (A).

Bottom Line: The stability of γD-crystallin against heat- or GdnHCl-induced denaturation was significantly decreased by the mutation, while no influence was observed on the acid-induced unfolding.The aggregation-prone property of the mutant was not altered by the addition of reductive reagent.These results suggested that the decrease in protein stability followed by aggregation-prone property might be the major cause in the hereditary cataract induced by the G61C mutation.

View Article: PubMed Central - PubMed

Affiliation: State Key Laboratory of Biomembrane and Membrane Biotechnology, School of Life Sciences, Tsinghua University, Beijing, China.

ABSTRACT
γD-crystallin is one of the major structural proteins in human eye lens. The solubility and stability of γD-crystallin play a crucial role in maintaining the optical properties of the lens during the life span of an individual. Previous study has shown that the inherited mutation G61C results in autosomal dominant congenital cataract. In this research, we studied the effects of the G61C mutation on γD-crystallin structure, stability and aggregation via biophysical methods. CD, intrinsic and extrinsic fluorescence spectroscopy indicated that the G61C mutation did not affect the native structure of γD-crystallin. The stability of γD-crystallin against heat- or GdnHCl-induced denaturation was significantly decreased by the mutation, while no influence was observed on the acid-induced unfolding. The mutation mainly affected the transition from the native state to the intermediate but not that from the intermediate to the unfolded or aggregated states. At high temperatures, both proteins were able to form aggregates, and the aggregation of the mutant was much more serious than the wild type protein at the same temperature. At body temperature and acidic conditions, the mutant was more prone to form amyloid-like fibrils. The aggregation-prone property of the mutant was not altered by the addition of reductive reagent. These results suggested that the decrease in protein stability followed by aggregation-prone property might be the major cause in the hereditary cataract induced by the G61C mutation.

Show MeSH
Related in: MedlinePlus