April 2009
Volume 50, Issue 13
Free
ARVO Annual Meeting Abstract  |   April 2009
First Low Resolution Structural Evidence of Full-length TGFBIp and Its Implications in TGFBI-linked Corneal Dystrophy
Author Affiliations & Notes
  • R. V. Basaiawmoit
    Center for Insoluble Protein Structures (inSPIN) and Interdisciplinary Nanoscience Center (iNANO), University of Aarhus, Aarhus, Denmark
  • C. L. P. Oliveira
    Department of Chemistry, University of Aarhus, Aarhus, Denmark
  • K. Runager
    Center for Insoluble Protein Structures (inSPIN) and Interdisciplinary Nanoscience Center (iNANO), University of Aarhus, Aarhus, Denmark
  • T. Kristensen
    Center for Insoluble Protein Structures (inSPIN) and Interdisciplinary Nanoscience Center (iNANO), University of Aarhus, Aarhus, Denmark
  • J. J. Enghild
    Center for Insoluble Protein Structures (inSPIN) and Interdisciplinary Nanoscience Center (iNANO), University of Aarhus, Aarhus, Denmark
  • J. S. Pedersen
    Department of Chemistry, University of Aarhus, Aarhus, Denmark
  • D. E. Otzen
    Center for Insoluble Protein Structures (inSPIN) and Interdisciplinary Nanoscience Center (iNANO), University of Aarhus, Aarhus, Denmark
  • Footnotes
    Commercial Relationships  R.V. Basaiawmoit, None; C.L.P. Oliveira, None; K. Runager, None; T. Kristensen, None; J.J. Enghild, None; J.S. Pedersen, None; D.E. Otzen, None.
  • Footnotes
    Support  inSPIN Post-doctoral grant, NEI Grant R01EY2712
Investigative Ophthalmology & Visual Science April 2009, Vol.50, 4311. doi:
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      R. V. Basaiawmoit, C. L. P. Oliveira, K. Runager, T. Kristensen, J. J. Enghild, J. S. Pedersen, D. E. Otzen; First Low Resolution Structural Evidence of Full-length TGFBIp and Its Implications in TGFBI-linked Corneal Dystrophy. Invest. Ophthalmol. Vis. Sci. 2009;50(13):4311.

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Abstract

Purpose: : TGFBI-linked corneal dystrophies are a rare hereditary condition characterised by proteinacious deposits in the corneal tissue which compromise corneal transparency and thus severely affect vision. It is now well known that the deposits mostly comprise of the protein Transforming Growth Factor Beta Induced Protein or TGFBIp in short (also referred to as keratoepithelin). Despite considerable interest a high resolution structure of the 683 residue protein has been lacking. Using Small Angle X-Ray Scattering and complementary biophysical and biochemical methods we have been able to determine the low resolution structure of wildtype (WT) TGFBIp and a corneal dystrophy mutant R124H.

Methods: : Small Angle Xray Scattering (SAXS), Size Exclusion Chromatography, Fluorescence Spectroscopy, Fluoresence Anisotropy, Dynamic Light Scattering (DLS), Chemical Cross linking, Native Polyacrylamide Gel Electrophoresis.

Results: : The structure of WT TGFBIp and R124H mutant revealed no structural differences, in the low resolution scale offered by SAXS. The protein exhibited real-time multimerisation properties which SAXS, DLS, Anisotropy and biochemical studies confirmed. The dominant species observed was calculated to be that of a trimer but both monomeric and dimeric species were also trapped and characterised.

Conclusions: : The low resolution structure for full length TGFBIp opens the door to understanding the molecular basis of Corneal Dystrophies. Furthermore, we suggest that the multimerisation properties seen for full length TGFBIp are probably linked to the multi-functional nature of the protein. No apparent structural differences, within the resolution offered by SAXS, between the WT and the R124H mutant (an aggressive corneal dystrophy mutant) thus suggest that the mutation must affect other cellular interactions to cause the patho-physiology known for this mutant.

Keywords: cornea: basic science • extracellular matrix • protein structure/function 
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