ADP-ribosylation of actins by arginine-specific ADP-ribosyltransferase purified from chicken heterophils.
We reported the purification and characterization of an arginine-specific ADP-ribosyltransferase and acceptor protein p33 in granules of chicken peripheral polymorphonuclear leukocytes (heterophils) [Mishima, K., Terashima, M., Obara, S., Yamada, K., Imai, K. & Shimoyama, M. (1991) J. Biochem. (Tokyo) 110, 388-394]. In the present study, we obtained evidence that chicken non-muscle beta/gamma-actin, skeletal muscle alpha-actin and smooth-muscle gamma-actin were ADP ribosylated by the heterophil ADP-ribosyltransferase. The stoichiometry of ADP-ribose incorporation into these actins was 1.2 mol, 1.0 mol and 2.0 mol ADP-ribose/mol of beta/gamma-actin, alpha-actin and gamma-actin, respectively. The optimal pH for the ADP ribosylation was at pH 8.5, with the respective actin. Km values for NAD were calculated to be 30 microM with beta/gamma-actin, 35 microM with alpha-actin and 20 microM with gamma-actin. The Km values for the actin isoforms were 15 microM for beta/gamma-actin, 2.5 microM for alpha-actin and 10 microM for gamma-actin. ADP ribosylation of actin inhibited its capacity to polymerize, as determined by the increase in fluorescence intensity with N-(1-pyrenyl)iodoacetamide-labelled actin. Filamentous actin (F-actin) polymerized with the respective actin isoform was also ADP ribosylated, although the extent of the modification of F-actin was lower than that of globular actin (G-actin). In situ ADP ribosylation of beta/gamma-actin was evidenced with chicken peripheral heterophils permeabilized with saponin. Thus, the endogenous ADP ribosylation of actin in the heterophils may be involved in the cellular processes such as phagocytosis, secretion and migration.
Terashima
M
M
Department of Biochemistry, Shimane Medical University, Izumo, Japan.
Mishima
K
K
Yamada
K
K
Tsuchiya
M
M
Wakutani
T
T
Shimoyama
M
M
D000199:Actins / Q000378:metabolism*; D000246:Adenosine Diphosphate Ribose / Q000378:metabolism*; D000818:Animals; D001120:Arginine / Q000378:metabolism*; D002645:Chickens / Q000097:blood*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008970:Molecular Weight; D009504:Neutrophils / Q000201:enzymology*; D011065:Poly(ADP-ribose) Polymerases / Q000097:blood*
D000199:Actins; D000246:Adenosine Diphosphate Ribose; D001120:Arginine; D011065:Poly(ADP-ribose) Polymerases