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D. Aroca-Aguilar, F. Sánchez-Sánchez, S. Ghosh, M. Coca-Prados, J. Escribano; Recombinant Myocilin Interacts With the Matricellular Protein SPARC. Invest. Ophthalmol. Vis. Sci. 2010;51(13):6093.
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© ARVO (1962-2015); The Authors (2016-present)
Myocilin is an extracellular glycoprotein with unknown function which is associated with different types of glaucoma. We have recently reported a proteolytical processing of myocilin by Calpain II within the linker region of the protein releasing the C-terminal olfactomedin domain. We have described also that myocilin interacts with the C-terminal region of hevin, a secretory glycoprotein belonging to the SPARC family of matricellular proteins. The main objective of this study is to investigate the interaction of recombinant myocilin with SPARC.
Analysis of protein-protein interactions were performed using the yeast two-hybrid system and solid-phase binding assays with Ni-chelating HPLC purified recombinant proteins. Coexpression of myocilin, SPARC and hevin in ocular tissues was confirmed by immunoflorescence microscopy, western blot and array-based gene profiling.
Yeast two-hybrid analyses showed that myocilin interacted with the highly conserved C-terminal extracellular calcium binding (EC) domain within SPARC and hevin. Solid-phase binding assays showed that full-length myocilin interacted with higher affinity with SPARC and its EC domain than the myocilin C-terminal fragment.
Recombinant myocilin and SPARC interact through their C-terminal domains. Our data suggest that the proteolytic processing of myocilin modulates this interaction supporting a functional role for this proteolytic clevage.
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