Residues within the polycationic region of cGMP phosphodiesterase gamma subunit crucial for the interaction with transducin alpha subunit. Identification by endogenous ADP-ribosylation and site-directed mutagenesis.

The Journal of Biological Chemistry
V A BondarenkoA Yamazaki

Abstract

Interaction between the gamma subunit (Pgamma) of cGMP phosphodiesterase and the alpha subunit (Talpha) of transducin is a key step for the regulation of cGMP phosphodiesterase in retinal rod outer segments. Here we have utilized a combination of specific modification by an endogenous enzyme and site-directed mutagenesis of the Pgamma polycationic region to identify residues required for the interaction with Talpha. Pgamma, free or complexed with the alphabeta subunit (Palphabeta) of cGMP phosphodiesterase, was specifically radiolabeled by prewashed rod membranes in the presence of [adenylate-32P]NAD. Identification of ADP-ribose in the radiolabeled Pgamma and radiolabeling of arginine-replaced mutant forms of Pgamma indicate that both arginine 33 and arginine 36 are similarly ADP-ribosylated by endogenous ADP-ribosyltransferase, but only one arginine is modified at a time. Pgamma complexed with Talpha (both GTP- and GDP-bound forms) was not ADP-ribosylated; however, agmatine, which cannot interact with Talpha, was ADP-ribosylated in the presence of Talpha, suggesting that a Pgamma domain containing these arginines is masked by Talpha. A Pgamma mutant (R33,36K), as well as wild type Pgamma, inhibited both GTP hydrolysis of Talp...Continue Reading

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Jan 30, 2002·The Journal of General Physiology·Peter D CalvertClint L Makino
Sep 9, 2006·Microbiology and Molecular Biology Reviews : MMBR·Paul O HassaMichael O Hottiger
Jun 6, 2014·Nature Reviews. Cancer·Sejal Vyas, Paul Chang
Sep 24, 2004·Neurochemistry International·John W Wellard, Ian G Morgan
Mar 25, 2017·International Journal of Molecular Sciences·Eric P KnottDamien D Pearse
Dec 14, 2001·The Journal of Biological Chemistry·Michael J PagliaRick H Cote

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