ADP‐ribosylation of actins by arginine‐specific ADP‐ribosyltransferase purified from chicken heterophils

Abstract

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 β/γ‐actin, skeletal muscle α‐actin and smooth‐muscle γ‐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 β/γ‐actin, α‐actin and γ‐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 μM with β/γ‐actin, 35 μM with α‐actin and 20 μM with γ‐actin. The Km values for the actin isoforms were 15 μM for β/γ‐actin, 2.5 μM for α‐actin and 10 μM for γ‐actin. ADP ribosylation of actin inhibited its capacity to polymerize, as determined by the increase in fluoresence 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 β/γ‐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. Copyright © 1992, Wiley Blackwell. All rights reserved