Production and characterization of monoclonal antibodies specific for the functional domains of poly(ADP-ribose) polymerase.
Production and characterization of monoclonal antibodies specific for the functional domains of poly(ADP-ribose) polymerase.
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聚（ADP-核糖）聚合酶功能域特异性单克隆抗体的生产和表征。
DOI：
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发表时间：
1986
期刊：
影响因子：
--
通讯作者：
G. Poirier
中科院分区：
文献类型：
--
作者：
D. Lamarre;B. Talbot;Y. Leduc;S. Muller;G. Poirier
针对聚（ADP-核糖）聚合酶开发单克隆抗体，并分析其对NAD+和DNA结合片段的反应性。进行两次融合以获得杂交瘤，并通过表征其免疫球蛋白轻链来进一步筛选所得的抗聚（ADP-核糖）聚合酶抗体。分离出五种不同的杂交瘤，它们产生不同的免疫球蛋白轻链，所有这些轻链都对聚（ADP-核糖）聚合酶具有特异性。这些抗体的特异性，通过免疫印迹法对纯化的聚（ADP-核糖）聚合酶，其自降解片段，和通过有限的蛋白水解与胰凝乳蛋白酶和木瓜蛋白酶制备的片段。这些片段分别含有NAD+结合位点、DNA结合位点和自修饰位点。所有的单克隆抗体与对应于纯化的酶的116 kdalton（kDa）条带反应。四种抗体仅与含有DNA结合位点的46-kDa片段上的抗原位点反应。第五种抗体仅与74-和54-kDa片段上的明显不同的抗原位点反应，所述片段具有NAD+（底物）结合位点。纯化酶的主要自降解产物（69-和46-kDa片段）的免疫反应性证实了两组抗体之间的这种差异。对应于自身修饰位点的22-kDa片段不显示与抗体的任何免疫反应性。
Monoclonal antibodies were developed against poly(ADP-ribose) polymerase and analyzed for their reactivity against the NAD+- and DNA-binding fragments. Two fusions were performed to obtain hybridomas and the resulting anti-poly(ADP-ribose) polymerase antibodies were further screened by characterization of their immunoglobulin light chains. Five different hybridomas were isolated which produced different immunoglobulin light chains, all of which were specific for poly(ADP-ribose) polymerase. The specificities of these antibodies were determined by immunoblotting against the purified poly(ADP-ribose) polymerase, its autodegradation fragments, and the fragments prepared by limited proteolysis with chymotrypsin and papain. These fragments have been suggested to contain the NAD+-binding site, the DNA-binding site, and the automodification site, respectively. All the monoclonal antibodies reacted with the 116 kdalton (kDa) band corresponding to the purified enzyme. Four antibodies reacted exclusively with antigenic site(s) on the 46-kDa fragment which contains the DNA-binding site. A fifth antibody reacted exclusively with a clearly different antigenic site on the 74- and 54-kDa fragments which possess the NAD+ (substrate) binding site. The immunoreactivity with the major autodegradation products (69- and 46-kDa fragments) of the purified enzyme confirms this difference between the two groups of antibodies. The 22-kDa fragment corresponding to the auto-modification site does not show any immunoreactivity with the antibodies.
影响因子：
2.9
作者：
KAUFMANN, SH;EWING, CM;SHAPER, JH
通讯作者：
SHAPER, JH