April 2011
Volume 52, Issue 14
ARVO Annual Meeting Abstract  |   April 2011
Myocilin, Q-snare Homology And Membrane-associated Binding Partners
Author Affiliations & Notes
  • William M. Dismuke
    University of Arizona, Tucson, Arizona
  • Brian S. McKay
    University of Arizona, Tucson, Arizona
  • W Daniel Stamer
    Ophthalmology & Vision Science,
    University of Arizona, Tucson, Arizona
  • Footnotes
    Commercial Relationships  William M. Dismuke, None; Brian S. McKay, None; W Daniel Stamer, None
  • Footnotes
    Support  Unrestricted grant from Research to Prevent Blindness, NEI grant EY0112797 and the Interdisciplinary Training in Cardiovascular Research grant HL07249
Investigative Ophthalmology & Visual Science April 2011, Vol.52, 2068. doi:
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      William M. Dismuke, Brian S. McKay, W Daniel Stamer; Myocilin, Q-snare Homology And Membrane-associated Binding Partners. Invest. Ophthalmol. Vis. Sci. 2011;52(14):2068.

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      © ARVO (1962-2015); The Authors (2016-present)

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Purpose: : Mutations in the widely expressed protein myocilin result in a specific phenotype, ocular hypertension, glaucoma and blindness, yet the function of myocilin remains controversial and elusive. Our preliminary data suggest that membrane-associated myocilin is present in a large protein complex. Here we test the hypothesis that myocilin is a component of a SNARE complex.

Methods: : We first used sequence analysis to examine structural homology of myocilin to known SNARE components. Second we used five complementary biochemical methods to examine the hydrodynamic properties of the myocilin complex; rotary shadowing, velocity gradient sedimentation, gel exclusion chromatography, immunoprecipitation and western blotting.

Results: : Our sequence analysis of the coiled-coil domain in a number of mammals revealed a segment with a hydrophobic stripe that exhibits homology to highly conserved regions of known Q-SNAREs. We found myocilin to exist in two cellular pools when separated by PAGE; as a 55/57kDa monomer in the soluble fraction and as a large, SDS-resistant protein complex in the membrane fraction. From the membrane fraction, velocity gradient sedimentation showed myocilin separating as small dimer (6.4s) and a large complex (17.3s). Myocilin was detected in a complex separating at 405-440 kDa by gel exclusion chromatography. Immunoprecipitation showed an association of myocilin with another coiled-coil containing SNARE protein, VAMP 1.

Conclusions: : Our data demonstrates that membrane-associated myocilin is present in a large, globular, detergent-resistant protein complex similar to other SNARE complexes (Söllner T, et al. Nature, 1993). The Q-SNARE homology region identified within the coiled-coil may facilitate the interaction of myocilin with its binding partners (Chapman ER, et al. JBC 1994).

Keywords: anterior segment • protein structure/function • proteins encoded by disease genes 

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