Purification of PDE6 isozymes from mammalian retina.
Abstract
Abstract
The photoreceptor phosphodiesterase (PDE6) is the central effector of visual transduction in vertebrate retinal photoreceptors. Distinct isozymes of PDE6 exist in rods and cones. Mammalian retina serves as an abundant source of tissue for PDE6 purification. Methods are described for the isolation and purification of membrane-associated PDE6 from rod outer segment membranes. Purification of cone PDE6 from the soluble fraction of retinal extracts is also described. Several procedures that can purify the rod and cone isozymes to homogeneity, including anion exchange, hydrophobic interaction, gel filtration, hydroxyapatite, and immunoaffinity chromatography, are presented. A method to activate PDE6 by limited proteolysis of its inhibitory gamma-subunit is also provided.
Department
Molecular, Cellular and Biomedical Sciences
Publication Date
2005
Journal Title
Methods in Molecular Biology
Publisher
Humana Press
Digital Object Identifier (DOI)
10.1385/1-59259-839-0:125
Document Type
Book Chapter
Recommended Citation
Pentia, D.C., Hosier, S., Collupy, R.A., Valeriani, B.A., Cote, R.H. "Purification of PDE6 isozymes from mammalian retina." Phosphodiesterase Methods and Protocols. Claire Luginer, (Ed). Series: Methods in molecular biology. Volume 307. Clifton, N.J. : Humana Press, 2005. pp. 125-140. doi:10.1385/1-59259-839-0:125