Background: Sorsby’s fundus dystrophy (SFD) is caused by mutations in tissue inhibitor of metalloproteinase (TIMP)-3 and, with the exception of early onset, is similar to age-related macular degeneration. The pathological features of this condition relate to the accumulation of TIMP-3 in Bruch’s membrane.
Aims: To compare the extracellular membrane-binding characteristics of wild-type and four SFD-mutant TIMP-3s.
Methods: COS-7 cells were transfected with wild-type, Ser-181, Gly-167, Ser-156 and Tyr-168 SFD-mutant TIMP-3 cDNA. The TIMP-3 proteins subsequently synthesised were harvested, analysed by sodium dodecyl sulphate-polyacrylamide gel electrophoresis, semiquantified by ELISA and used in binding assays on the basis of the retention of the wild-type and SFD-mutant TIMP-3 proteins by components of Bruch’s membrane.
Results: SFD-mutant TIMP-3s could not be distinguished from wild-type TIMP-3 by the extents to which they aggregated or adhered to type-I collagen, type-IV collagen, fibronectin, laminin, elastin, chondroitin sulphates A, B and C, and heparin sulphate. Of these macromolecules, the wild-type and SFD-mutant TIMP-3s exhibited greatest affinity for elastin and laminin.
Conclusion: The similarity in the physical and extracellular membrane-binding characteristics of wild-type and SFD-mutant TIMP-3s indicates that these properties are not responsible for the difference in timing of onset of SFD and age-related macular degeneration.
- ARMD, age-related macular degeneration
- DMEM, Dulbecco’s modified Eagle medium
- ECM, extracellular membrane
- FCS, fetal calf serum
- GAG, glycosaminoglycan
- HRP, horseradish peroxidase
- MMP, matrix metalloproteinase
- RPE, retinal pigment epithelial
- SDS, sodium dodecyl sulphate
- SFD, Sorsby’s fundus dystrophy
- TIMP-3, tissue inhibitor of metalloproteinase-3
- TMB, 3,3′,5,5′-tetramethyl benzidine
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