Clancy R M, Leszczynska-Piziak J, Abramson S B
Department of Medicine, NYU Medical Center, Hospital for Joint Diseases, New York 10003.
Biochem Biophys Res Commun. 1993 Mar 31;191(3):847-52. doi: 10.1006/bbrc.1993.1294.
ADP-ribosylation is an important post-translational protein modification; however, endogenous substrates are poorly characterized. In these studies we examined the effects of nitric oxide on the ADP-ribosylation of neutrophil proteins. Purified cytosol and plasma membrane were incubated with 32P-NAD (5 microM, 1 microCi, 30 min) in the presence or absence of nitric oxide. Nitric oxide induced the ADP-ribosylation of the 37 kD substrate present only in cytosol. Nitric oxide treatment of plasma membrane plus cytosol revealed the ADP-ribosylation of an additional 43 kD protein. This 43 kD substrate was identified as actin by both phalloidin precipitation and immunoblot (2-D) gel using specific anti-actin antibodies. The data indicate that nitric oxide stimulates the ADP-ribosylation of two discrete substrates in fractionated PMN, one of which can be identified as actin. NO-induced ADP-ribosylation may contribute to the modulatory effect of nitric oxide on neutrophil functions, including F-actin assembly.
ADP核糖基化是一种重要的翻译后蛋白质修饰；然而，内源性底物的特征却鲜为人知。在这些研究中，我们检测了一氧化氮对中性粒细胞蛋白质ADP核糖基化的影响。在有或没有一氧化氮存在的情况下，将纯化的胞质溶胶和质膜与32P-NAD（5微摩尔，1微居里，30分钟）一起孵育。一氧化氮诱导了仅存在于胞质溶胶中的37kD底物的ADP核糖基化。对质膜加胞质溶胶进行一氧化氮处理后，发现了另一种43kD蛋白质的ADP核糖基化。通过鬼笔环肽沉淀和使用特异性抗肌动蛋白抗体的免疫印迹（二维）凝胶分析，将这种43kD底物鉴定为肌动蛋白。数据表明，一氧化氮刺激了分离的多形核白细胞中两种不同底物的ADP核糖基化，其中一种可鉴定为肌动蛋白。一氧化氮诱导的ADP核糖基化可能有助于一氧化氮对中性粒细胞功能（包括F-肌动蛋白组装）的调节作用。
