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Purification of PDE6 Isozymes From Mammalian Retina

  • Dana C. Pentia
  • Suzanne Hosier
  • Rachel A. Collupy
  • Beverly A. Valeriani
  • Rick H. Cote
Part of the Methods In Molecular Biology™ book series (MIMB, volume 307)

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 γ-subunit is also provided.

Key Words

Photoreceptor phosphodiesterase retina phototransduction anion-exchange chromatography cone and rod hydrophobic interaction chromatography 

Notes

Acknowledgment

This work was supported by National Eye Institute (National Institues of Health) grant EY 05798 and is Scientific Contribution Number 2195 from the New Hampshire Agricultural Experiment Station.

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Copyright information

© Humana Press Inc. 2005

Authors and Affiliations

  • Dana C. Pentia
    • 1
  • Suzanne Hosier
    • 1
  • Rachel A. Collupy
    • 1
  • Beverly A. Valeriani
    • 1
  • Rick H. Cote
    • 1
  1. 1.Department of Biochemistry and Molecular BiologyUniversity of New HampshireDurham

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