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Thermal stress induced aggregation of aquaporin 0 (AQP0) and protection by ?-crystallin via its chaperone function.

ABSTRACT: Aquaporin 0 (AQP0) formerly known as membrane intrinsic protein (MIP), is expressed exclusively in the lens during terminal differentiation of fiber cells. AQP0 plays an important role not only in the regulation of water content but also in cell-to-cell adhesion of the lens fiber cells. We have investigated the thermal stress-induced structural alterations of detergent (octyl glucoside)-solubilized calf lens AQP0. The results show an increase in the amount of AQP0 that aggregated as the temperature increased from 40°C to 65°C. ?-Crystallin, molecular chaperone abundantly present in the eye lens, completely prevented the AQP0 aggregation at a 1?1 (weight/weight) ratio. Since ?-crystallin consists of two gene products namely ?A- and ?B-crystallins, we have tested the recombinant proteins on their ability to prevent thermal-stress induced AQP0 aggregation. In contrast to the general observation made with other target proteins, ?A-crystallin exhibited better chaperone-like activity towards AQP0 compared to ?B-crystallin. Neither post-translational modifications (glycation) nor C-terminus truncation of AQP0 have any appreciable effect on its thermal aggregation properties. ?-Crystallin offers similar protection against thermal aggregation as in the case of the unmodified AQP0, suggesting that ?crystallin may bind to either intracellular loops or other residues of AQP0 that become exposed during thermal stress. Far-UV circular dichroism studies indicated a loss of ?helical structures when AQP0 was subjected to temperatures above 45°C, and the presence of ?-crystallin stabilized these secondary structures. We report here, for the first time, that ?-crystallin protects AQP0 from thermal aggregation. Since stress-induced structural perturbations of AQP0 may affect the integrity of the lens, presence of the molecular chaperone, ?-crystallin (particularly ?A-crystallin) in close proximity to the lens membrane is physiologically relevant.

SUBMITTER: Swamy-Mruthinti S

PROVIDER: S-EPMC3842347 | biostudies-literature | 2013

REPOSITORIES: biostudies-literature

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Thermal stress induced aggregation of aquaporin 0 (AQP0) and protection by α-crystallin via its chaperone function.

Swamy-Mruthinti Satyanarayana S Srinivas Volety V Hansen John E JE Rao Ch Mohan ChM

PloS one 20131127 11

Aquaporin 0 (AQP0) formerly known as membrane intrinsic protein (MIP), is expressed exclusively in the lens during terminal differentiation of fiber cells. AQP0 plays an important role not only in the regulation of water content but also in cell-to-cell adhesion of the lens fiber cells. We have investigated the thermal stress-induced structural alterations of detergent (octyl glucoside)-solubilized calf lens AQP0. The results show an increase in the amount of AQP0 that aggregated as the temperat ...[more]

PMID: 24312215

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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.

Dataset Information

Thermal stress induced aggregation of aquaporin 0 (AQP0) and protection by ?-crystallin via its chaperone function.

Publications

Thermal stress induced aggregation of aquaporin 0 (AQP0) and protection by α-crystallin via its chaperone function.

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