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Matrix bound SFD mutant TIMP-3 is more stable than wild type TIMP-3
  1. Mohammed A Majid1,
  2. Valerie A Smith1,
  3. Andrew C Newby2,
  4. Andrew D Dick1
  1. 1Academic unit of Ophthalmology, University of Bristol, Bristol Eye Hospital, Bristol, UK
  2. 2Bristol Heart Institute, University of Bristol, Bristol Royal Infirmary, Bristol, UK
  1. Correspondence to: Valerie A Smith Bristol Eye Hospital, Lower Maudlin Street, Bristol BS1 2LX, UK; Val.Smith{at}bristol.ac.uk

Abstract

Background: Sorsby’s fundus dystrophy (SFD) is a degenerative retinopathy characterised by accumulation of mutant TIMP-3 protein in Bruch’s membrane.

Aim: To compare the stability of matrix bound SFD mutant TIMP-3s with wild type TIMP-3.

Methods: COS-7 cells were transfected with plasmids containing wild type, Ser 181, Gly-167, Ser-156, and Tyr-168 TIMP-3 cDNA. The cells and their matrices were subsequently harvested and homogenised. After measuring the bound wild type and SFD mutant TIMP-3 concentrations by ELISA, aliquots of the homogenates were heated to 100°C. The rates of denaturation of the TIMP proteins at this temperature were monitored by reverse zymography.

Results: Over a period of 24 h at 100°C the biological activity of both wild type and SFD mutant TIMP-3 was lost. Over a period of 6 h at this temperature the biological activity of the SFD mutant TIMP-3s was fully retained whereas that of the wild type TIMP-3 was lost.

Conclusion: Matrix bound SFD mutant TIMP-3s are thermodynamically more stable than wild type. This may explain why SFD starts earlier in life than age related macular degeneration.

  • ARMD, age related macular degeneration
  • ELISA, enzyme linked immunosorbent assay
  • RPE, retinal pigment epithelial
  • SFD, Sorsby’s fundus dystrophy
  • Bruch’s membrane
  • TIMP-3 gene transfer
  • Sorsby’s fundus dystrophy
  • age related macular degeneration
  • ARMD, age related macular degeneration
  • ELISA, enzyme linked immunosorbent assay
  • RPE, retinal pigment epithelial
  • SFD, Sorsby’s fundus dystrophy
  • Bruch’s membrane
  • TIMP-3 gene transfer
  • Sorsby’s fundus dystrophy
  • age related macular degeneration

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Footnotes

  • Published Online First 23 March 2007

  • Competing interests: None declared.

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