April 2010
Volume 51, Issue 13
Free
ARVO Annual Meeting Abstract  |   April 2010
Disease Mutations in the Second Extracellular Loop of the Human Retina Specific ABC Transporter, ABCA4, Impart Structural Defects in This Domain
Author Affiliations & Notes
  • E. E. Biswas-Fiss
    Bioscience Technologies, Thomas Jefferson University, Philadelphia, Pennsylvania
  • D. S. Kurpad
    Bioscience Technologies, Thomas Jefferson University, Philadelphia, Pennsylvania
  • K. Joshi
    Bioscience Technologies, Thomas Jefferson University, Philadelphia, Pennsylvania
  • B. Sajer
    Bioscience Technologies, Thomas Jefferson University, Philadelphia, Pennsylvania
  • Footnotes
    Commercial Relationships  E.E. Biswas-Fiss, None; D.S. Kurpad, None; K. Joshi, None; B. Sajer, None.
  • Footnotes
    Support  NIH Grant EY 013113
Investigative Ophthalmology & Visual Science April 2010, Vol.51, 1093. doi:https://doi.org/
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      E. E. Biswas-Fiss, D. S. Kurpad, K. Joshi, B. Sajer; Disease Mutations in the Second Extracellular Loop of the Human Retina Specific ABC Transporter, ABCA4, Impart Structural Defects in This Domain. Invest. Ophthalmol. Vis. Sci. 2010;51(13):1093. doi: https://doi.org/.

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

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Abstract

Purpose: : The retina specific ABC transporter, ABCA4, is associated with a broad range of inherited macular degenerations including Stargardt disease (STDG), autosomal recessive cone rod dystrophy (arCRD) and fundus flavimaculatus (FFM). A unique topological feature of the ABCA family of ABC transporters is the presence of long extracellular (EC) loops or domains, sequences of which are highly conserved. A cluster of point mutations in the second extracellular domain (ECD2) has been identified in patients with the aforementioned degenerative diseases.

Methods: : We have analyzed the solution structure of this domain by circular dichroism (CD) spectroscopy and investigated the structural changes upon single, disease- associated point mutations.

Results: : CD analysis of purified, recombinant forms of this protein demonstrated that the ECD2 domain has a highly ordered and stable structure comprised of 27±3% alpha helix, 20±3% beta-pleated sheet, and 53±3% coil. Significant changes in the CD spectra were observed in disease mutants as well as in their predicted protein conformations. Thermal melting by CD spectroscopic analysis demonstrated alteration in the overall stability of the structures of ECD2 upon mutation. The point mutations also altered the environment in the core of the protein structure as observed through measurement of intrinsic tryptophan fluorescence (EX 295 nm and EM of 320-500 nm).

Conclusions: : These results support the hypothesis that the extracellular domains of ABCA4 play an important role in its transport function which is likely dependent upon specific conformational features of these domains; furthermore Stargardt disease associated mutations may lead to significant changes in the structural features of these domains.

Keywords: protein structure/function • retinal degenerations: hereditary • proteins encoded by disease genes 
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