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[63] Purification of rod outer segment GTP-Binding protein subunits and cgmp phosphodiesterase by single-step column chromatography

Authors
Publisher
Elsevier Science & Technology
Identifiers
DOI: 10.1016/0076-6879(88)59065-1
Keywords
  • Section Iv. Methods For Isolation And Studies Of Various Posppdoesterase Isoenzyme B. Cgmp-Binding P
Disciplines
  • Biology
  • Physics

Abstract

Publisher Summary This chapter describes a novel and simple single-step chromatographic method for the extensive purification of the subunits of the GTP-binding protein and the phosphodiesterase (PDE). There is a well-established homology between the hormone-sensitive adenylate cyclase and the light-activated cGMP phosphodiesterase in rod outer segments (ROS). The light-activated PDE of ROS has three functional segments. The photopigment rhodopsin, the GTP-binding protein, and the PDE catalytic complex. Signal transduction from rhodopsin to PDE is dependent on the binding of GTP to the α subunit of the GTP-binding protein. Photon capture is necessary for the binding of GTP to the Gα subunit. The GTP/Gα complex is rapidly released from the disk membrane surface. The activation of PDE in ROS prepared from the amphibian retina occurs as a consequence of the presentation of photons and the addition of GTP. Moreover, this activation depends upon the physical release of an inhibitory moiety from the PDE that remains bound to the disk membrane throughout the activation cycle. The major components of the ROS light-sensitive PDE cascade have been isolated and purified. The purification of rhodopsin has been well characterized in a number of laboratories.

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