Dataset Information

Acetylation of Gly1 and Lys2 promotes aggregation of human ?D-crystallin.

ABSTRACT: The human lens contains three major protein families: ?-, ?-, and ?-crystallin. Among the several variants of ?-crystallin in the human lens, ?D-crystallin is a major form. ?D-Crystallin is primarily present in the nuclear region of the lens and contains a single lysine residue at the second position (K2). In this study, we investigated the acetylation of K2 in ?D-crystallin in aging and cataractous human lenses. Our results indicated that K2 is acetylated at an early age and that the amount of K2-acetylated ?D-crystallin increased with age. Mass spectrometric analysis revealed that in addition to K2, glycine 1 (G1) was acetylated in ?D-crystallin from human lenses and in ?D-crystallin acetylated in vitro. The chaperone ability of ?-crystallin for acetylated ?D-crystallin was lower than that for the nonacetylated protein. The tertiary structure and the microenvironment of the cysteine residues were significantly altered by acetylation. The acetylated protein exhibited higher surface hydrophobicity, was unstable against thermal and chemical denaturation, and exhibited a higher propensity to aggregate at 80 °C in comparison to the nonacetylated protein. Acetylation enhanced the GdnHCl-induced unfolding and slowed the subsequent refolding of ?D-crystallin. Theoretical analysis indicated that the acetylation of K2 and G1 reduced the structural stability of the protein and brought the distal cysteine residues (C18 and C78) into close proximity. Collectively, these results indicate that the acetylation of G1 and K2 residues in ?D-crystallin likely induced a molten globule-like structure, predisposing it to aggregation, which may account for the high content of aggregated proteins in the nucleus of aged and cataractous human lenses.

SUBMITTER: DiMauro MA

PROVIDER: S-EPMC4245984 | biostudies-literature | 2014 Nov

REPOSITORIES: biostudies-literature

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Acetylation of Gly1 and Lys2 promotes aggregation of human γD-crystallin.

DiMauro Michael A MA Nandi Sandip K SK Raghavan Cibin T CT Kar Rajiv Kumar RK Wang Benlian B Bhunia Anirban A Nagaraj Ram H RH Biswas Ashis A

Biochemistry 20141113 46

The human lens contains three major protein families: α-, β-, and γ-crystallin. Among the several variants of γ-crystallin in the human lens, γD-crystallin is a major form. γD-Crystallin is primarily present in the nuclear region of the lens and contains a single lysine residue at the second position (K2). In this study, we investigated the acetylation of K2 in γD-crystallin in aging and cataractous human lenses. Our results indicated that K2 is acetylated at an early age and that the amount of ...[more]

PMID: 25393041

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Project description:Cataract, a major cause of visual impairment worldwide, is the opacification of the eye's crystalline lens due to aggregation of the crystallin proteins. The research reported here is aimed at investigating the aggregating behavior of γ-crystallin proteins in various incubation conditions. Thioflavin T binding assay, circular dichroism spectroscopy, 1-anilinonaphthalene-8-sulfonic acid fluorescence spectroscopy, intrinsic (tryptophan) fluorescence spectroscopy, light scattering, and electron microscopy were used for structural characterization. Molecular dynamics simulations and bioinformatics prediction were performed to gain insights into the γD-crystallin mechanisms of fibrillogenesis. We first demonstrated that, except at pH 7.0 and 37°C, the aggregation of γD-crystallin was observed to be augmented upon incubation, as revealed by turbidity measurements. Next, the types of aggregates (fibrillar or non-fibrillar aggregates) formed under different incubation conditions were identified. We found that, while a variety of non-fibrillar, granular species were detected in the sample incubated under pH 7.0, the fibrillogenesis of human γD-crystallin could be induced by acidic pH (pH 2.0). In addition, circular dichroism spectroscopy, 1-anilinonaphthalene-8-sulfonic acid fluorescence spectroscopy, and intrinsic fluorescence spectroscopy were used to characterize the structural and conformational features in different incubation conditions. Our results suggested that incubation under acidic condition led to a considerable change in the secondary structure and an enhancement in solvent-exposure of the hydrophobic regions of human γD-crystallin. Finally, molecular dynamics simulations and bioinformatics prediction were performed to better explain the differences between the structures and/or conformations of the human γD-crystallin samples and to reveal potential key protein region involved in the varied aggregation behavior. Bioinformatics analyses revealed that the initiation of amyloid formation of human γD-crystallin may be associated with a region within the C-terminal domain. We believe the results from this research may contribute to a better understanding of the possible mechanisms underlying the pathogenesis of senile nuclear cataract.

Dataset Information

Acetylation of Gly1 and Lys2 promotes aggregation of human ?D-crystallin.

Publications

Acetylation of Gly1 and Lys2 promotes aggregation of human γD-crystallin.

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