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Structural features of isomerizable aspartyl residues in human ?-crystallins.

ABSTRACT: PURPOSE: The aspartyl (Asp) residues 58 and 151 in ?A-crystallin, and Asp36 and Asp62 in ?B-crystallin in human lenses are known to be highly isomerized with aging. We investigate structural environments of these isomerizable aspartyl residues in ?-crystallins of human lenses. METHODS: To perform limited proteolysis experiments of purified human ?A- and ?B-crystallins, endoproteinase Asp-N (EC 3.4.24.33), which selectively cleaves the peptide bonds at the amino side of aspartyl and cysteic acid residues, was employed. By proteolysis approach coupled with the time-of-flight mass spectrometry (TOF-MS) method, we determined the cleavage points along protein sequences. RESULTS: Proteolysis by endoproteinase Asp-N occurred preferentially at the site of isomerizable aspartyl residues in ?A- and ?B-crystallins. CONCLUSIONS: It is found that isomerizable aspartyl residues in ?-crystallins in human lenses were located not only in the solvent accessible area but also at regions displaying inherent conformational flexibility.

SUBMITTER: Shimizu K

PROVIDER: S-EPMC3398496 | biostudies-literature | 2012

REPOSITORIES: biostudies-literature

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Structural features of isomerizable aspartyl residues in human α-crystallins.

Shimizu Ken-ichi K Kita Akiko A Fujii Noriko N Miki Kunio K

Molecular vision 20120704

<h4>Purpose</h4>The aspartyl (Asp) residues 58 and 151 in αA-crystallin, and Asp36 and Asp62 in αB-crystallin in human lenses are known to be highly isomerized with aging. We investigate structural environments of these isomerizable aspartyl residues in α-crystallins of human lenses.<h4>Methods</h4>To perform limited proteolysis experiments of purified human αA- and αB-crystallins, endoproteinase Asp-N (EC 3.4.24.33), which selectively cleaves the peptide bonds at the amino side of aspartyl and ...[more]

PMID: 22815635

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Project description:Crystallins are transparent, refractive proteins that contribute to the focusing power of the vertebrate eye lens. These proteins are extremely soluble and resist aggregation for decades, even under crowded conditions. Crystallins have evolved to avoid strong interprotein interactions and have unusual hydration properties. Crystallin aggregation resulting from mutation, damage, or aging can lead to cataract, a disease state characterized by opacity of the lens.Different aggregation mechanisms can occur, following multiple pathways and leading to aggregates with varied morphologies. Studies of variant proteins found in individuals with childhood-onset cataract have provided insight into the molecular factors underlying crystallin stability and solubility. Modulation of exposed hydrophobic surface is critical, as is preventing specific intermolecular interactions that could provide nucleation sites for aggregation. Biophysical measurements and structural biology techniques are beginning to provide a detailed picture of how crystallins crowd into the lens, providing high refractivity while avoiding excessively tight binding that would lead to aggregation.Despite the central biological importance of refractivity, relatively few experimental measurements have been made for lens crystallins. Our work and that of others have shown that hydration is important to the high refractive index of crystallin proteins, as are interactions between pairs of aromatic residues and potentially other specific structural features.This Account describes our efforts to understand both the functional and disease states of vertebrate eye lens crystallins, particularly the ?-crystallins. We use a variety of biophysical techniques, notably NMR spectroscopy, to investigate crystallin stability and solubility. In the first section, we describe efforts to understand the relative stability and aggregation propensity of different ?S-crystallin variants. The second section focuses on interactions of these proteins with the holdase chaperone ?B-crystallin. The third, fourth, and fifth sections explore different modes of aggregation available to crystallin proteins, and the final section highlights the importance of refractive index and the sometimes conflicting demands of selection for refractivity and solubility.

Dataset Information

Structural features of isomerizable aspartyl residues in human ?-crystallins.

Publications

Structural features of isomerizable aspartyl residues in human α-crystallins.

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