June 2015
Volume 56, Issue 7
ARVO Annual Meeting Abstract  |   June 2015
Interaction of human gammaS-crystallin and its cataract-associated G18V mutant with alpha-crystallins
Author Affiliations & Notes
  • Ajay Pande
    Chemistry, University at Albany, Albany, NY
  • Jayanti Pande
    Chemistry, University at Albany, Albany, NY
  • Footnotes
    Commercial Relationships Ajay Pande, None; Jayanti Pande, None
  • Footnotes
    Support None
Investigative Ophthalmology & Visual Science June 2015, Vol.56, 5593. doi:
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      Ajay Pande, Jayanti Pande; Interaction of human gammaS-crystallin and its cataract-associated G18V mutant with alpha-crystallins. Invest. Ophthalmol. Vis. Sci. 2015;56(7 ):5593.

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

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Most gamma crystallins bind to alpha crystallins under conditions favoring their unfolding, while some gamma crystallin variants in their native state also bind alpha crystallins. Here we explore the binding of human gammaS-crystallin and its cataract-associated mutant G18V to human alphaA-crystallin


Recombinant human gammaS- and alphaA- crystallins were expressed in E. coli. G18V was prepared by site-directed mutagenesis of gammaS-crystallin. Binding of gammaS (or mutant) under native or unfolding conditions in mixtures with alphaA was analyzed using size-exclusion chromatography with a Superose-6 column. The higher molecular weight fraction containing alphaA and its complex with gammaS (or mutant) was dialyzed using a 100K-cutoff ultrafiltration device. Hydrodynamic diameter was determined using dynamic light scattering. Spectral analysis was used to characterize different forms of binding.<br />


Preliminary studies indicate that gammaS-crystallin binds to alphaA-crystallin under thermal-unfolding conditions, like the other gamma-crystallins. The chromatographic protein fraction representing alphaA and its complex shows the spectral signature of the substrate gammaS-crysallin. Increasing amount of complex formation occurs as the duration of thermal stress increases, and the effective hydrodynamic diameter increases from ~19 nm for native alphaA to as high as 32 nm. Interestingly, it has been shown that gammaS does not bind to alphaB under similar conditions. We also find that G18V in the native state, does not bind to alphaA-crystallin. This contrasts with its interaction with alphaB-crystallin which it has been shown to bind. Thus alphaA- and alphaB-crystallins appear to behave differently with gammaS-crystallin and its G18V mutant.<br />


The interaction of human gammaS-crystallin with alphaA-crystallin appears to differ from that of alphaB-crystallin. We therefore conclude that the molecular chaperones, alphaA and alphaB-crystallins exhibit different specificities towards the same set of substrate or target proteins. Work is currently in progress to substantiate these findings.


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