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J. H. McDowell, D. R. Dugger, A. Arendt, S. Bolch, W. C. Smith; Interaction of Arrestin With Alpha Enolase in Photoreceptors. Invest. Ophthalmol. Vis. Sci. 2010;51(13):1095. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
Arrestin is in disequilibrium in photoreceptors, changing its partitioning between inner and outer segments in response to lighting conditions. The purpose of this project was to identify the cellular component with which arrestin associates in the dark-adapted retina.
Whole bovine retinas were cross-linked with 2.5 mM DSP, and arrestin-containing complexes were purified by anion exchange chromatography. Tandem mass spectrometry analysis was used to identify the protein components in the complex. Enolase1-specific antibodies were produced and used to assess enolase1 localization in photoreceptors by indirect fluorescent immunohistochemistry. Surface plasmon resonance (SPR) was used to measure direct interaction between enolase and arrestin. An enolase activity assay was used to assess the influence of arrestin on enolase activity.
In whole retina treated with DSP, arrestin crosslinks in a 125 kD complex. The principal components of this complex are arrestin and enolase1. In the dark-adapted retina, enolase1 co-localizes with arrestin in the inner segments and outer nuclear layer, but is also present in the outer plexiform layer. SPR analysis of purified arrestin and enolase1 demonstrates direct binding interaction between arrestin and enolase1. In an enolase activity assay, arrestin slows the catalytic activity of enolase by a maximum of 24%.
Our results show that in the dark-adapted retina, arrestin is in proximity to enolase 1, sufficiently close that it can be cross-linked by DSP. Our SPR data show that the interaction between arrestin and enolase is direct, not requiring a third element to form the complex. The presence of arrestin slows the catalytic activity of enolase, suggesting that light-driven translocation of arrestin may be a method to modulate the metabolic activity of photoreceptors.
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