Investigative ophthalmology & visual science · 2025
Identification of Age- and Cataract-Related Changes in High-Density Lens Protein Aggregates.
СтатьяПоддержка NIH: внешняя программаПоддержка исследования: вне правительства СШАНазвание и аннотация: Перевод пока не подготовлен
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Wang Zhen, Friedrich Michael G, Truscott Roger J W, Schey Kevin L
Аннотация
PURPOSE: Cataract is believed to be caused by protein-protein and protein-membrane aggregation in the eye lens. After middle age, there is extensive binding of crystallins to the lens cell membranes as evidenced by sedimentation at high densities. Multiple protein modifications have been linked with cataract, whereas others have been associated with aging. The purpose of this study was to characterize protein constituents within high density protein-membrane fractions from normal aged or cataractous lenses and to compare these proteins and their modifications.
METHODS: The inner nuclear regions of cataract or age-matched normal lenses were homogenized and proteins were separated using sucrose density gradient centrifugation. The low-density fractions (LDFs) and high-density fractions (HDFs) were analyzed by mass spectrometry using both top-down matrix-assisted desorption/ionization-mass spectrometry (MALDI-MS) and bottom-up liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based proteomic methods. Quantification of low molecular weight crystallin peptides, deamidation, and isomerization were performed.
RESULTS: Compared with normal aged-lenses, membrane-associated protein aggregates in high density fractions of cataract lenses exhibited significantly higher levels of γ-crystallins, as well as γS- and γD-crystallin C-terminal peptides. Deamidation of γ-crystallin, but not of β-crystallin, was increased in cataract lens membrane-bound aggregates. A very high level of Asp isomerization was detected in bound α-crystallins from both aged and cataract lenses.
CONCLUSIONS: Binding of crystallin aggregates to human lens cell membranes is associated with protein truncation, deamidation, and isomerization, and was observed in normal aged and cataract lenses. However, the protein aggregates bound to membranes in cataract lenses exhibit distinct modifications to γ-crystallins that may arise as a consequence of additional protein degradation.
METHODS: The inner nuclear regions of cataract or age-matched normal lenses were homogenized and proteins were separated using sucrose density gradient centrifugation. The low-density fractions (LDFs) and high-density fractions (HDFs) were analyzed by mass spectrometry using both top-down matrix-assisted desorption/ionization-mass spectrometry (MALDI-MS) and bottom-up liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based proteomic methods. Quantification of low molecular weight crystallin peptides, deamidation, and isomerization were performed.
RESULTS: Compared with normal aged-lenses, membrane-associated protein aggregates in high density fractions of cataract lenses exhibited significantly higher levels of γ-crystallins, as well as γS- and γD-crystallin C-terminal peptides. Deamidation of γ-crystallin, but not of β-crystallin, was increased in cataract lens membrane-bound aggregates. A very high level of Asp isomerization was detected in bound α-crystallins from both aged and cataract lenses.
CONCLUSIONS: Binding of crystallin aggregates to human lens cell membranes is associated with protein truncation, deamidation, and isomerization, and was observed in normal aged and cataract lenses. However, the protein aggregates bound to membranes in cataract lenses exhibit distinct modifications to γ-crystallins that may arise as a consequence of additional protein degradation.