Pharmacological chaperone for ?-crystallin partially restores transparency in cataract models.
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ABSTRACT:
SUBMITTER: Makley LN
PROVIDER: S-EPMC4725592 | biostudies-other | 2015 Nov
REPOSITORIES: biostudies-other
Publications
Pharmacological chaperone for α-crystallin partially restores transparency in cataract models.
Makley Leah N LN McMenimen Kathryn A KA DeVree Brian T BT Goldman Joshua W JW McGlasson Brittney N BN Rajagopal Ponni P Dunyak Bryan M BM McQuade Thomas J TJ Thompson Andrea D AD Sunahara Roger R Klevit Rachel E RE Andley Usha P UP Gestwicki Jason E JE
Science (New York, N.Y.) 20151101 6261
Cataracts reduce vision in 50% of individuals over 70 years of age and are a common form of blindness worldwide. Cataracts are caused when damage to the major lens crystallin proteins causes their misfolding and aggregation into insoluble amyloids. Using a thermal stability assay, we identified a class of molecules that bind α-crystallins (cryAA and cryAB) and reversed their aggregation in vitro. The most promising compound improved lens transparency in the R49C cryAA and R120G cryAB mouse model ...[more]
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Project description:BackgroundA substitution mutation in human ?A-crystallin (?AG98R) is associated with autosomal dominant cataract. The recombinant mutant ?AG98R protein exhibits altered structure, substrate-dependent chaperone activity, impaired oligomer stability and aggregation on prolonged incubation at 37 °C. Our previous studies have shown that ?A-crystallin-derived mini-chaperone (DFVIFLDVKHFSPEDLTVK) functions like a molecular chaperone by suppressing the aggregation of denaturing proteins. The present study was undertaken to determine the effect of ?A-crystallin-derived mini-chaperone on the stability and chaperone activity of ?AG98R-crystallin.Methodology/principal findingsRecombinant ?AG98R was incubated in presence and absence of mini-chaperone and analyzed by chromatographic and spectrometric methods. Transmission electron microscope was used to examine the effect of mini-chaperone on the aggregation propensity of mutant protein. Mini-chaperone containing photoactive benzoylphenylalanine was used to confirm the interaction of mini-chaperone with ?AG98R. The rescuing of chaperone activity in mutant?-crystallin (?AG98R) by mini-chaperone was confirmed by chaperone assays. We found that the addition of the mini-chaperone during incubation of ?AG98R protected the mutant crystallin from forming larger aggregates that precipitate with time. The mini-chaperone-stabilized ?AG98R displayed chaperone activity comparable to that of wild-type ?A-crystallin. The complexes formed between mini-?A-?AG98R complex and ADH were more stable than the complexes formed between ?AG98R and ADH. Western-blotting and mass spectrometry confirmed the binding of mini-chaperone to mutant crystallin.Conclusion/significanceThese results demonstrate that mini-chaperone stabilizes the mutant ?A-crystallin and modulates the chaperone activity of ?AG98R. These findings aid in our understanding of how to design peptide chaperones that can be used to stabilize mutant ?A-crystallins and preserve the chaperone function.
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