Abstract
Purpose: :
To analyze potential role of soy isoflavones as antioxidants in aggregation of crystallins in water soluble (WS) protein fractions of young and old monkey (Macaca fascicularis) lenses.
Methods: :
A total of twelve monkey lenses were divided into the following four groups for this study: (i) young control (6 to 9 year-old on soy-free diet), (ii) young on soy-diet (6 to 9 year-old fed soy [2 mg/g] for 30 months), (iii) old control (15 year-old or older on soy-free diet), and (iv) old on soy-diet (15-year-old or older fed soy [2 mg/g] for 30 months). The WS proteins from individual lenses in each group were examined by multi-angle light scattering (MALS) instrument to determine the molecular mass of high molecular weight (HMW) proteins and crystallin fractions. The WS protein fractions of each lens were also analyzed by 2D-gel electrophoresis and 2-D Fluorescence Difference Gel Electrophoresis (2-D DIGE). During 2-D DIGE, identical amounts of WS proteins from each lens were derivatized with Cy2, Cy3 and Cy5 fluorophores using the manufacturer’s protocol. Image analysis was done using DecyderTM Differential Analysis Software (GE Healthcare Life Sciences).
Results: :
The MALS results showed that the -crystallin peak was higher in the lenses of young monkeys on soy-diet compared to young controls on soy-free diet. The 2D-gel analysis of proteins from young monkey lenses on soy-diet and soy-free diets also showed differences in the β-crystallin region. Additionally, the 2D-gel results showed greater numbers of post translationally modified -crystallin species and truncated crystallins in older monkey lenses on soy-free diet compared to those on soy-diet. At five-fold lower concentrations of the WS proteins, the 2-D DIGE analyses showed differences in 4 spots in young control monkey lenses vs. young lenses on soy-diet. However, similar 2-D DIGE analyses showed no significant changes in protein profiles of lenses from older monkeys on soy-diet or soy-free diet.
Conclusions: :
The results show increased truncation and aggregation of crystallins in older monkey lenses on soy-free diet compared to those on soy-diet. Apparently, soy-diet was more effective in protection of -crystallin in young lenses.
Keywords: crystallins • protein modifications-post translational • antioxidants