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αA-Crystallin–Derived Mini-Chaperone Modulates Stability and Function of Cataract Causing αAG98R-Crystallin
BACKGROUND: A substitution mutation in human αA-crystallin (αAG98R) is associated with autosomal dominant cataract. The recombinant mutant αAG98R protein exhibits altered structure, substrate-dependent chaperone activity, impaired oligomer stability and aggregation on prolonged incubation at 37°C. O...
Autores principales: | , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Public Library of Science
2012
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3435407/ https://www.ncbi.nlm.nih.gov/pubmed/22970163 http://dx.doi.org/10.1371/journal.pone.0044077 |
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author | Raju, Murugesan Santhoshkumar, Puttur Sharma, K. Krishna |
author_facet | Raju, Murugesan Santhoshkumar, Puttur Sharma, K. Krishna |
author_sort | Raju, Murugesan |
collection | PubMed |
description | BACKGROUND: A substitution mutation in human αA-crystallin (αAG98R) is associated with autosomal dominant cataract. The recombinant mutant αAG98R 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 αA-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 αA-crystallin–derived mini-chaperone on the stability and chaperone activity of αAG98R-crystallin. METHODOLOGY/PRINCIPAL FINDINGS: Recombinant αAG98R 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 αAG98R. The rescuing of chaperone activity in mutantα-crystallin (αAG98R) by mini-chaperone was confirmed by chaperone assays. We found that the addition of the mini-chaperone during incubation of αAG98R protected the mutant crystallin from forming larger aggregates that precipitate with time. The mini-chaperone-stabilized αAG98R displayed chaperone activity comparable to that of wild-type αA-crystallin. The complexes formed between mini-αA–αAG98R complex and ADH were more stable than the complexes formed between αAG98R and ADH. Western-blotting and mass spectrometry confirmed the binding of mini-chaperone to mutant crystallin. CONCLUSION/SIGNIFICANCE: These results demonstrate that mini-chaperone stabilizes the mutant αA-crystallin and modulates the chaperone activity of αAG98R. These findings aid in our understanding of how to design peptide chaperones that can be used to stabilize mutant αA-crystallins and preserve the chaperone function. |
format | Online Article Text |
id | pubmed-3435407 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2012 |
publisher | Public Library of Science |
record_format | MEDLINE/PubMed |
spelling | pubmed-34354072012-09-11 αA-Crystallin–Derived Mini-Chaperone Modulates Stability and Function of Cataract Causing αAG98R-Crystallin Raju, Murugesan Santhoshkumar, Puttur Sharma, K. Krishna PLoS One Research Article BACKGROUND: A substitution mutation in human αA-crystallin (αAG98R) is associated with autosomal dominant cataract. The recombinant mutant αAG98R 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 αA-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 αA-crystallin–derived mini-chaperone on the stability and chaperone activity of αAG98R-crystallin. METHODOLOGY/PRINCIPAL FINDINGS: Recombinant αAG98R 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 αAG98R. The rescuing of chaperone activity in mutantα-crystallin (αAG98R) by mini-chaperone was confirmed by chaperone assays. We found that the addition of the mini-chaperone during incubation of αAG98R protected the mutant crystallin from forming larger aggregates that precipitate with time. The mini-chaperone-stabilized αAG98R displayed chaperone activity comparable to that of wild-type αA-crystallin. The complexes formed between mini-αA–αAG98R complex and ADH were more stable than the complexes formed between αAG98R and ADH. Western-blotting and mass spectrometry confirmed the binding of mini-chaperone to mutant crystallin. CONCLUSION/SIGNIFICANCE: These results demonstrate that mini-chaperone stabilizes the mutant αA-crystallin and modulates the chaperone activity of αAG98R. These findings aid in our understanding of how to design peptide chaperones that can be used to stabilize mutant αA-crystallins and preserve the chaperone function. Public Library of Science 2012-09-06 /pmc/articles/PMC3435407/ /pubmed/22970163 http://dx.doi.org/10.1371/journal.pone.0044077 Text en © 2012 Raju et al http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are properly credited. |
spellingShingle | Research Article Raju, Murugesan Santhoshkumar, Puttur Sharma, K. Krishna αA-Crystallin–Derived Mini-Chaperone Modulates Stability and Function of Cataract Causing αAG98R-Crystallin |
title | αA-Crystallin–Derived Mini-Chaperone Modulates Stability and Function of Cataract Causing αAG98R-Crystallin |
title_full | αA-Crystallin–Derived Mini-Chaperone Modulates Stability and Function of Cataract Causing αAG98R-Crystallin |
title_fullStr | αA-Crystallin–Derived Mini-Chaperone Modulates Stability and Function of Cataract Causing αAG98R-Crystallin |
title_full_unstemmed | αA-Crystallin–Derived Mini-Chaperone Modulates Stability and Function of Cataract Causing αAG98R-Crystallin |
title_short | αA-Crystallin–Derived Mini-Chaperone Modulates Stability and Function of Cataract Causing αAG98R-Crystallin |
title_sort | αa-crystallin–derived mini-chaperone modulates stability and function of cataract causing αag98r-crystallin |
topic | Research Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3435407/ https://www.ncbi.nlm.nih.gov/pubmed/22970163 http://dx.doi.org/10.1371/journal.pone.0044077 |
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