NADP/sup +/ enhances cholera and pertussis toxin-catalyzed ADP-ribosylation of membrane proteins

Conference · Thu May 01 00:00:00 EDT 1986 · Fed. Proc., Fed. Am. Soc. Exp. Biol.; (United States)

OSTI ID:5326819

Kawai, Y; Whitsel, C; Arinze, I J

Cholera or pertussis toxin-catalyzed (/sup 32/P)ADP-ribosylation is frequently used to estimate the concentration of the stimulatory (Ns) or inhibitory (Ni) guanine nucleotide regulatory proteins which modulate the activity of adenylate cyclase. With this assay, however, the degradation of the substrate, NAD/sup +/, by endogenous enzymes such as NAD/sup +/-glycohydrolase (NADase) present in the test membranes can influence the results. In this study the authors show that both cholera and pertussis toxin-catalyzed (/sup 32/P)ADP-ribosylation of liver membrane proteins is markedly enhanced by NADP/sup +/. The effect is concentration dependent; with 20 ..mu..M (/sup 32/P)NAD/sup +/ as substrate maximal enhancement is obtained at 0.5-1.0 mM NADP/sup +/. The enhancement of (/sup 32/P)ADP-ribosylation by NADP/sup +/ was much greater than that by other known effectors such as Mg/sup 2 +/, phosphate or isoniazid. The effect of NADP/sup +/ on ADP-ribosylation may occur by inhibition of the degradation of NAD/sup +/ probably by acting as an alternate substrate for NADase. Among inhibitors tested (NADP/sup +/, isoniazid, imidazole, nicotinamide, L-Arg-methyl-ester and HgCl/sub 2/) to suppress NADase activity, NADP/sup +/ was the most effective and, 10 mM, inhibited activity of the enzyme by about 90%. In membranes which contain substantial activities of NADase the inclusion of NADP/sup +/ in the assay is necessary to obtain maximal ADP-ribosylation.

• Research Organization: Meharry Medical College, Nashville, TN
• OSTI ID: 5326819
• Report Number(s): CONF-8606151-
• Journal Information: Fed. Proc., Fed. Am. Soc. Exp. Biol.; (United States), Journal Name: Fed. Proc., Fed. Am. Soc. Exp. Biol.; (United States) Vol. 45:6; ISSN FEPRA
• Country of Publication: United States
• Language: English

Related Subjects

550201* -- Biochemistry-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
ADP
ANTIGENS
BETA DECAY RADIOISOTOPES
BETA-MINUS DECAY RADIOISOTOPES
BIOCHEMICAL REACTION KINETICS
BODY
CATALYSTS
CELL CONSTITUENTS
CELL MEMBRANES
COENZYMES
CYCLASES
DAYS LIVING RADIOISOTOPES
DIGESTIVE SYSTEM
ENZYME ACTIVITY
ENZYME INHIBITORS
ENZYMES
GLANDS
ISOTOPE APPLICATIONS
ISOTOPES
KINETICS
LIGHT NUCLEI
LIVER
LYASES
MATERIALS
MEMBRANES
METABOLITES
NAD
NADP
NUCLEI
NUCLEOTIDES
ODD-ODD NUCLEI
ORGANIC COMPOUNDS
ORGANS
PHOSPHORUS 32
PHOSPHORUS ISOTOPES
PROTEINS
RADIOISOTOPES
REACTION KINETICS
TOXIC MATERIALS
TOXINS
TRACER TECHNIQUES