Purification and characterization of an osteoclast membrane glycoprotein with homology to manganese superoxide dismutase.
Purification and characterization of an osteoclast membrane glycoprotein with homology to manganese superoxide dismutase.
复制标题
与锰超氧化物歧化酶同源的破骨细胞膜糖蛋白的纯化和表征。
DOI：
10.1002/jcb.240460305
复制
发表时间：
1991
影响因子：
4
通讯作者：
Osdoby,P
中科院分区：
文献类型：
--
作者：
Oursler,MJ;Li,L;Osdoby,P
破骨细胞是专门的多核细胞，主要负责骨基质的无机和有机成分的降解。分离的禽类破骨细胞已用于免疫小鼠并产生破骨细胞定向的单克隆抗体文库（J. Cell Biology，100：1592）。这些单克隆抗体的一个子集识别在破骨细胞上表达的抗原，而在体外由单核细胞或骨髓单核细胞形成的多核巨细胞上不存在或几乎不存在。这些抗体之一，命名为121 F，已被用于鉴定和纯化破骨细胞质膜相关糖蛋白。对体外形成的破骨细胞或多核巨细胞的二硫键还原提取物的Western印迹分析表明，121 F抗体识别仅在破骨细胞中可检测到的150 kDa蛋白。这种高分子量蛋白质已通过免疫亲和和凝胶过滤色谱法结合SDS-聚丙烯酰胺凝胶电洗脱单一条带进行纯化。SDS-聚丙烯酰胺凝胶上纯化抗原的银染色显示，未还原形式的单个蛋白质分子大于200 kDa，二硫化物还原时大于150 kDa。纯化抗原的等电聚焦显示单一种类，具有6.95的中性pl点。内切糖苷酶处理和凝集素亲和色谱分析表明，121 F抗体识别的抗原具有复杂的N-连接糖，而三氟甲磺酸处理表明不存在额外的O-连接糖组分。高碘酸盐氧化和单糖半抗原抑制研究未提供121 F抗体结合的抗原表位本质上为碳水化合物的证据。尽管天然抗原在其N末端被封闭，但羟胺生成肽的氨基酸分析揭示了121 F单克隆抗体识别的破骨细胞抗原与锰和铁超氧化物歧化酶之间的惊人关系。因此，除了作为破骨细胞的细胞类型特异性标志物外，这种细胞表面糖蛋白可能直接在破骨细胞介导的骨吸收中发挥作用。
The osteoclast is the specialized multinucleated cell primarily responsible for the degradation of the inorganic and organic components of bone matrix. Isolated avian osteoclasts have been used to immunize mice and generate an osteoclast‐directed monoclonal antibody library (J. Cell Biology, 100:1592). A subset of these monoclonal antibodies recognizes antigens which are expressed on osteoclasts and which are absent or nearly so on multinucleated giant cells formed in vitro from monocyte or marrow mononuclear cells. One of these antibodies, designated 121F, has been used to identify and purify an osteoclast plasma membrane‐associated glycoprotein. Western blot analysis on disulfide bond‐reduced extracts from osteoclasts or multinucleated giant cells formed in vitro demonstrates that the 121F antibody recognizes a 150 kDa protein detectable only in osteoclasts. This high molecular weight protein has been purified by a combination of immunoaffinity and gel filtration chromatography procedures, in conjunction with electroelution of a single band from SDS‐polyacrylamide gels. Silver staining of the purified antigen on SDS‐polyacrylamide gels has revealed a single protein species larger than 200 kDa in its unreduced form and 150 kDa when disulfides are reduced. Isoelectric focusing of the purified antigen reveals a single species, having a neutral pl point of 6.95. Whereas endoglycosidase treatment and lectin affinity chromatographic analyses demonstrate that the antigen recognized by the 121F antibody possesses complex N‐linked sugars, trifluoromethanesulfonic acid treatment indicates there are no additional O‐linked carbohydrate components. Periodate oxidation and monosaccharide hapten inhibition studies provide no evidence for the antigenic epitope bound by the 121F antibody being carbohydrate in nature. Although the native antigen is blocked at its N‐terminus, amino acid analysis of a hydroxylamine generated peptide disclosed a striking relationship between the osteoclast antigen recognized by the 121F monoclonal antibody and manganese and iron superoxide dismutase. Therefore, in addition to serving as a distinguishing cell type–specific marker for osteoclasts, this cell surface glycoprotein may function directly in osteoclast‐mediated bone resorption.
登录
查看更多内容
千里光使动物中毒。
DOI：
--
发表时间：
1934
期刊：
影响因子：
--
作者：
D. Steyn
通讯作者：
D. Steyn
服用吡咯里西啶生物碱的大鼠的急性肝毒性和吡咯代谢物。
影响因子：
5.1
作者：
A. Mattocks
通讯作者：
A. Mattocks
体外肝微粒体由吡咯里西啶生物碱形成的吡咯和氮氧化物代谢物：与体内肝毒性的相关性。
影响因子：
5.1
作者：
A. Mattocks;I. Bird
通讯作者：
I. Bird
DOI：
--
发表时间：
1989-07
期刊：
Drug metabolism and disposition: the biological fate of chemicals
影响因子：
--
作者：
David E. Williams;Ralph L. Reed;Bogdan Kedzierski;G. Dannan;F. Guengerich;Donald R. Buhler
通讯作者：
David E. Williams;Ralph L. Reed;Bogdan Kedzierski;G. Dannan;F. Guengerich;Donald R. Buhler
影响因子：
3.8
作者：
GREEN, CE;SEGALL, HJ;BYARD, JL
通讯作者：
BYARD, JL