April 2009
Volume 50, Issue 13
ARVO Annual Meeting Abstract  |   April 2009
Lens Crystallin Aggregation: The Tale of Two A-Crystallin-Derived Peptides
Author Affiliations & Notes
  • K. Sharma
    Ophthalmology and Biochemistry,
    University of Missouri, Columbia, Missouri
  • P. Santhoshkumar
    University of Missouri, Columbia, Missouri
  • R. Murugesan
    University of Missouri, Columbia, Missouri
  • Footnotes
    Commercial Relationships  K. Sharma, None; P. Santhoshkumar, None; R. Murugesan, None.
  • Footnotes
    Support  NIH Grants EY11981, EY14795, EY09855 and an Unrestricted grant to the department of Ophthalmology from Research to Prevent Blindness (RPB)
Investigative Ophthalmology & Visual Science April 2009, Vol.50, 2105. doi:
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      K. Sharma, P. Santhoshkumar, R. Murugesan; Lens Crystallin Aggregation: The Tale of Two A-Crystallin-Derived Peptides. Invest. Ophthalmol. Vis. Sci. 2009;50(13):2105.

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      © ARVO (1962-2015); The Authors (2016-present)

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Purpose: : Recently we identified over 25 low molecular mass (<3.5-kDa) peptides in the urea-soluble fractions of aged and cataract human lenses. (J. Biol. Chem. Vol 283, 8477-8485, 2008). Among the peptides identified several were from A-crystallin chaperone site or subunit interaction region. This study was undertaken to determine the role of two crystallin fragments - A66-80 and A67-75 found in protein aggregation in lenses.

Methods: : The levels of A66-80 and A67-75 in human lens were estimated by Mass spectrometric method. The conformation and amyloid forming propensity of in vitro synthesized A66-80 and A67-75 peptides was determined by CD spectroscopy and TEM whereas the Congo-red and Thioflavin-T dye binding assays were used to further confirm the amyloid peptide features. The anti-chaperone and -crystallin aggregation activities of the two peptides were measured by aggregation assays.

Results: : We found that a 70+ yr lens water-insoluble fraction has significantly higher amount of peptides than the water-soluble fraction. We estimated that lens has 98 ± 25 ng of A67-75 and 1.123 ± 0.23 µg of A66-80. Spectroscopic studies of A-66-80 peptide showed β-sheet conformation. On incubation at 37oC in 7.2 pH buffer both A-66-80 and A67-75 peptides formed amyloid fibril-like structures. There was also a time-dependent increase in the fluorescence of Congo Red and Thioflavin-T dyes in presence of A-66-80 or A67-75 peptide. Additionally, the interaction of A-66-80 or A67-75 peptide with -crystallin diminished the chaperone activity of the latter. The light scattering at 37oC by denaturing alcohol dehydrogenase was enhanced by A-66-80 and A67-75 peptides. Further, co-incubation of A-66-80 or A67-75 peptide with human lens extract resulted in aggregation and precipitation of -crystallin.

Conclusions: : Our study shows that A-crystallin-derived A-66-80 and A67-75 peptides present in human lenses form amyloid-like structures, interact with other crystallins, induce protein aggregation and show anti-chaperone activities. Therefore, the accumulation of these crystallin fragments in vivo and their interaction with native and modified crystallins may be an early event in cataractogenesis.

Keywords: crystallins • chaperones • proteolysis 

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