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The congenital cataract-linked A2V mutation impairs tetramer formation and promotes aggregation of ?B2-crystallin.

ABSTRACT: ?/?-Crystallins, the major structural proteins in human lens, are highly conserved in their tertiary structures but distinct in the quaternary structures. The N- and C-terminal extensions have been proposed to play a crucial role in mediating the size of ?-crystallin assembly. In this research, we investigated the molecular mechanism underlying the congenital hereditary cataract caused by the recently characterized A2V mutation in ?B2-crystallin. Spectroscopic experiments indicated that the mutation did not affect the secondary and tertiary structures of ?B2-crystallin. The mutation did not affect the formation of ?B2/?A3-crystallin heteromer as well as the stability and folding of the heteromer, suggesting that the mutation might not interfere with the protein interacting network in the lens. However, the tetramerization of ?B2-crystallin at high protein concentrations was retarded by the A2V mutation. The mutation slightly decreased the thermal stability and promoted the thermal aggregation of ?B2-crystallin. Although it did not influence the stability of ?B2-crystallin against denaturation induced by chemical denaturants and UV irradiation, the A2V mutant was more prone to be trapped in the off-pathway aggregation process during kinetic refolding. Our results suggested that the A2V mutation might lead to injury of lens optical properties by decreasing ?B2-crystallin stability against heat treatment and by impairing ?B2-crystallin assembly into high-order homo-oligomers.

SUBMITTER: Xu J

PROVIDER: S-EPMC3516508 | biostudies-literature | 2012

REPOSITORIES: biostudies-literature

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The congenital cataract-linked A2V mutation impairs tetramer formation and promotes aggregation of βB2-crystallin.

Xu Jia J Wang Sha S Zhao Wei-Jie WJ Xi Yi-Bo YB Yan Yong-Bin YB Yao Ke K

PloS one 20121206 12

β/γ-Crystallins, the major structural proteins in human lens, are highly conserved in their tertiary structures but distinct in the quaternary structures. The N- and C-terminal extensions have been proposed to play a crucial role in mediating the size of β-crystallin assembly. In this research, we investigated the molecular mechanism underlying the congenital hereditary cataract caused by the recently characterized A2V mutation in βB2-crystallin. Spectroscopic experiments indicated that the muta ...[more]

PMID: 23236454

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Project description:BackgroundHuman γS-crystallin is an important component of the human eye lens nucleus and cortex. The mutation V42M in the molecule causes severe congenital cataract in children. We compare the structure of the mutant protein with that of the wild type in order to understand how structural changes in the mutant relate to the mechanism of opacification.MethodsBoth proteins were made using conventional cloning and expression procedures. Secondary and tertiary structural features of the proteins were analyzed using spectral methods. Structural stabilities of the proteins were analyzed using chemical and thermal denaturation methods. Self-aggregation was monitored using extrinsic spectral probes. Molecular modeling was used to compare the structural features of the two proteins.ResultsWhile the wild type and mutant have the same secondary structure, molecular modeling and fluorescence analysis suggest the mutant to have a more open tertiary structure, with a larger hydrophobic surface. Experiments using extrinsic probes reveal that the mutant readily self-aggregates, with the suggestion that the aggregates might be similar to amyloidogenic fibrils. Chemical denaturation indicates that while the wild type exhibits the classic two-state transition, V42M goes through an intermediate state, and has a distinctly lower stability than the wild type. The temperature of thermal unfolding of the mutant is also distinctly lower. Further, the mutant readily precipitates and scatters light more easily than the wild type.ConclusionThe replacement of valine in position 42 by the longer and bulkier methionine in human γS-crystallin perturbs the compact β-sheet core packing topology in the N-terminal domain of the molecule, exposes nonpolar residues thereby increasing the surface hydrophobicity and weakens the stability of the protein, thus promoting self-aggregation leading to light scattering particles. This set of changes in the properties of the mutant offers a molecular insight into the mechanism of opacification.

Dataset Information

The congenital cataract-linked A2V mutation impairs tetramer formation and promotes aggregation of ?B2-crystallin.

Publications

The congenital cataract-linked A2V mutation impairs tetramer formation and promotes aggregation of βB2-crystallin.

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