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Y. V. Sergeev, J. F. Hejtmancik; Apoferritin Is Maintaining the Native Conformation of Citrate Synthase in vitro. Invest. Ophthalmol. Vis. Sci. 2009;50(13):3402.
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Ferritin, a member of family of iron storage proteins, is found in a wide variety of different human tissues including cornea, retina and RPE. Ferritin has been shown to increase in response to stress and inflammation and to sequester iron-II in the cell. In the eye rising levels of ferritin were detected in hyper-ferritinemia cataract and age-related macular degeneration. Ferritin expression under oxidative or temperature stress conditions and its protective effect on cell viability suggest that ferritin lacking iron (apoferritin) may have a role in the formation, stabilization or maintenance of the native conformation of proteins.
To test this hypothesis we studied the influence of apoferritin on the unfolding and refolding of citrate synthase (CS) in vitro using aggregation assay, size exclusion chromatography, SDS-PAGE and Western blot analysis, and fluorescent spectroscopy. The native state of CS was determined by measuring catalytic activity.
Here we show that at stoichiometric amounts apoferritin protects CS catalytic activity, stabilize the aggregation of CS under heat stress,and acts as a chaperone-like molecule in these folding reactions in vitro. Furthermore, apoferritin promotes the functional refolding of CS after guanidinium hydrochloride denaturation. Experiments on biotin label transfer demonstrated that iron-free apoferritin participates in transient protein-protein interactions with protein substrates, whereas ferritin does not show this interaction.
These results confirm that apoferritin has chaperone-like activity in vitro and suggests that in addition to its iron binding activity apoferritin might have a role in protecting and maintaining the native conformation of proteins in the eye.
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