BIOCHEMICAL AND FUNCTIONAL SIMILARITIES BETWEEN HUMAN EOSINOPHIL-DERIVED NEUROTOXIN AND EOSINOPHIL CATIONIC PROTEIN - HOMOLOGY WITH RIBONUCLEASE
BIOCHEMICAL AND FUNCTIONAL SIMILARITIES BETWEEN HUMAN EOSINOPHIL-DERIVED NEUROTOXIN AND EOSINOPHIL CATIONIC PROTEIN - HOMOLOGY WITH RIBONUCLEASE
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DOI：
10.1073/pnas.83.10.3146
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发表时间：
1986-05-01
影响因子：
11.1
通讯作者：
MCKEAN, DJ
中科院分区：
文献类型：
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作者：
GLEICH, GJ;LOEGERING, DA;MCKEAN, DJ
用凝胶过滤和肝素-琼脂糖离子交换层析从人嗜酸性粒细胞颗粒裂解液中分离出嗜酸性粒细胞源性神经毒素（EDN）和嗜酸性粒细胞阳离子蛋白（ECP）。使用单克隆抗体对来自肝素-琼脂糖层析的组分进行放射免疫测定，显示EDN活性的一个峰和ECP活性的两个峰（称为ECP-1和ECP-2）。EDN，ECP-1和ECP-2的电荷和分子量的异质性，当通过二维非平衡pH梯度电泳和NaDodSO 4/PAGE分析。用内切糖苷酶F（endoF）消化EDN降低了其分子量和电荷异质性。因此，EDN可能含有单一复合寡糖。ECP-1和ECP-2的Endo F消化降低了两种多肽的分子量，表明两者可能含有至少一种复合寡糖。氨基酸序列分析表明，ECP-1和ECP-2从残基1至残基59是相同的，EDN和ECP的序列是高度同源的（55个残基中有37个相同）。EDN和ECP的NH 2-末端序列显示出显着的同源性RNase，特别是在区域的RNase分子参与配体结合。EDN、ECP-1和ECP-2具有神经毒性活性，当注射到小脑延髓池中时，在低至0.15 μ g的剂量下引起戈登现象;这些蛋白质的活性相当。这些结果表明，EDN和ECP是相关的蛋白质，并表明它们来自与RNase家族相关的基因。
Eosinophil-derived neurotoxin (EDN) and eosinophil cationic protein (ECP) were isolated from lysates of human eosinophil granules by gel filtration and ion exchange chromatography on heparin-Sepharose. Radioimmunoassay, using monoclonal antibodies, of fractions from the heparin-Sepharose chromatography showed one peak of EDN activity and two peaks of ECP activity (termed ECP-1 and ECP-2). EDN, ECP-1, and ECP-2 each exhibited heterogeneity in charge and molecular weight when analyzed by two-dimensional nonequilibrium pH gradient electrophoresis and NaDodSO4/PAGE. Digestion of EDN with endoglycosidase F (endoF) decreased its molecular weight and charge heterogeneity. Thus, EDN likely contains a single complex oligosaccharide. Endo F digestion of ECP-1 and ECP-2 decreased the molecular weight of both polypeptides, indicating that both likely contain at least one complex oligosaccharide. Amino acid sequence analyses showed that ECP-1 and ECP-2 are identical from residue 1 through residue 59 and that the sequences of EDN and ECP are highly homologous (37 of 55 residues identical). Both EDN and ECP NH2-terminal sequences showed significant homology to RNase, especially in regions of the RNase molecule involved in ligand binding. EDN, ECP-1, and ECP-2 had neurotoxic activity, causing the Gordon phenomenon at doses down to 0.15 .mu.g when injected into the cisterna magna; the proteins were comparable in their activities. These results indicate that EDN and ECP are related proteins and suggest that they derived from genes associated with the RNase family.