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Om P. Srivastava, Kiran Srivastava, Jose M. Chaves; Human Lens Crystallin Fragments Form Aggregates Alone And With Water Soluble-High Molecular Weight (HMW)-Proteins That Also Exist In Vivo. Invest. Ophthalmol. Vis. Sci. 2011;52(14):4747.
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© ARVO (1962-2015); The Authors (2016-present)
The purpose was to determine aggregation of crystallin fragments alone and with WS-HMW proteins and whether these aggregates exist in vivo.
The WS-protein fraction from lenses of 50-70 year-old human donors was fractionated by a preparative SDS-PAGE method to isolate following four fractions containing crystallin fragments: Fraction I (3.5 kDa species), Fraction II (3.5 to 7 kDa), fraction III (7 to10 kDa) and Fraction IV (>10-18 kDa). Following 2D-gel electrophoresis, individual spots in Fraction I to IV were identified by a mass spectrometric method. The self-aggregation of crystallin fragments in the above four fractions and with WS-HMW proteins was determined by a size-exclusion HPLC method.
The 2D-gel electrophoretic profiles showed 13 spots in Fraction I, and 36, 46 and 53 spots in Fractions II, III and IV, respectively. The Q-TRAP mass spectrometric analysis showed that individual spots contained mutiple fragments of α-, β- and γ-crystallins with a variety of post-translational modifications.The fragments of Fractions I to IV exhibited self-aggregtaion on storage to species with molecular mass between 1.5X105 to1.5X107 Da, .and also with WS-HMW proteins. Following mass spectrometric analysis of 2-D electrophoretically-separated spots of WS-HMW proteins, the presence of crystalllin fragments as complexes was observed.
Crystallin fragments of α-, β- and γ-crystallins with a variety of post-translational modifications exist in aging humam lenses. Their presence in WS-HMW proteins, and properties of self-aggregation per se and with WS-HMW proteins suggest a potential role during aggregation process in vivo.
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