The bacterial toxins, choleragen and pertussis toxin, inhibit the light-stimulated GTPase activity of bovine retinal rod outer segments by catalysing the ADP-ribosylation of the alpha-subunit (T alpha) of transducin [Abood, Hurley, Pappone, Bourne & Stryer (1982) J. Biol. Chem. 257, 10540-10543; Van Dop, Yamanaka, Steinberg, Sekura, Manclark, Stryer & Bourne (1984) J. Biol. Chem. 259, 23-26]. Incubation of retinal rod outer segments with NAD+ and a purified NAD+:arginine ADP-ribosyltransferase from turkey erythrocytes resulted in approx. 60% inhibition of GTPase activity. Inhibition was dependent on both enzyme and NAD+, and was potentiated by the non-hydrolysable GTP analogues guanosine 5′-[beta gamma-imido]triphosphate (p[NH]ppG) and guanosine 5′-[beta gamma-methylene]triphosphate (p[CH2]ppG). The transferase ADP-ribosylated both the T alpha and T beta subunits of purified transducin. T alpha (39 kDa), after ADP-ribosylation, migrated as two distinct peptides with molecular masses of 42 kDa and 46 kDa on SDS/polyacrylamide-gel electrophoresis. T beta (36 kDa), after ADP-ribosylation, migrated as a 38 kDa peptide. With purified transducin subunits, it was observed that the GTPase activity of ADP-ribosylated T alpha, reconstituted with unmodified T beta gamma and photolysed rhodopsin, was decreased by 80%; conversely, reconstitution of T alpha with ADP-ribosyl-T beta gamma resulted in only a 19% inhibition of GTPase. Thus ADP-ribosylation of T alpha, the transducin subunit that contains the guanine nucleotide-binding site, has more dramatic effects on GTPase activity than does modification of the critical ‘helper subunits’ T beta gamma. To elucidate the mechanism of GTPase inhibition by transferase, we studied the effect of ADP-ribosylation on p[NH]pp[3H]G binding to transducin. It was shown previously that modification of transducin by choleragen, which like transferase ADP-ribosylates arginine residues, did not affect guanine nucleotide binding. ADP-ribosylation by the transferase, however, decreased p[NH]pp[3H]G binding, consistent with the hypothesis that choleragen and transferase inhibit GTPase by different mechanisms.
Skip Nav Destination
Follow us on Twitter @Biochem_Journal
Article navigation
December 1987
-
Cover Image
Cover Image
- PDF Icon PDF LinkFront Matter
- PDF Icon PDF LinkTable of Contents
- PDF Icon PDF LinkAdvertising
Research Article|
December 15 1987
Inhibition of the GTPase activity of transducin by an NAD+:arginine ADP-ribosyltransferase from turkey erythrocytes
P A Watkins;
P A Watkins
Laboratory of Cellular Metabolism, National Heart, Lung, and Blood Institute, National Insititutes of Health, Bethesda, MD 20892, U.S.A.
Search for other works by this author on:
Y Kanaho;
Y Kanaho
Laboratory of Cellular Metabolism, National Heart, Lung, and Blood Institute, National Insititutes of Health, Bethesda, MD 20892, U.S.A.
Search for other works by this author on:
J Moss
J Moss
Laboratory of Cellular Metabolism, National Heart, Lung, and Blood Institute, National Insititutes of Health, Bethesda, MD 20892, U.S.A.
Search for other works by this author on:
Publisher: Portland Press Ltd
Online ISSN: 1470-8728
Print ISSN: 0264-6021
© 1987 London: The Biochemical Society
1987
Biochem J (1987) 248 (3): 749–754.
Citation
P A Watkins, Y Kanaho, J Moss; Inhibition of the GTPase activity of transducin by an NAD+:arginine ADP-ribosyltransferase from turkey erythrocytes. Biochem J 15 December 1987; 248 (3): 749–754. doi: https://doi.org/10.1042/bj2480749
Download citation file:
Sign in
Don't already have an account? Register
Sign in to your personal account
You could not be signed in. Please check your email address / username and password and try again.
Could not validate captcha. Please try again.
Biochemical Society Member Sign in
Sign InSign in via your Institution
Sign in via your InstitutionGet Access To This Article
Follow us on Twitter @Biochem_Journal
Open Access for all
We offer compliant routes for all authors from 2025. With library support, there will be no author nor reader charges in 5 journals. Check here |
![]() View past webinars > |