Exposure on cell surface and extensive arginine methylation of ewing sarcoma (EWS) protein
Exposure on cell surface and extensive arginine methylation of ewing sarcoma (EWS) protein
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DOI：
10.1074/jbc.m011446200
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发表时间：
2001-06-01
影响因子：
4.8
通讯作者：
Gehring, H
中科院分区：
文献类型：
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作者：
Belyanskaya, LL;Gehrig, PM;Gehring, H
与有关染色体易位引起的嵌合ewing肉瘤（EWS）融合蛋白功能的知识相反，RNA结合EWS蛋白的细胞功能的表征很差。 EWS蛋白主要在细胞核中发现。在本报告中，我们表明EWS蛋白不仅在细胞核和细胞质中发现，而且在细胞中也发现了EWS蛋白。表面。在细胞表面生物素化后，膜分数的等电聚焦，生物素基化蛋白的抗原蛋白 - 琼脂糖提取以及SDS-聚丙烯酰胺凝胶电泳，EWS蛋白通过基质辅助激光脱离电离电离和纳米素质质量质量群鉴定。消化的肽。这些分析表明，具有重复的RGG基序的蛋白质在精氨酸残基上广泛非对称二甲基化，其位点已通过质谱方法映射。在总共30个Arg-Gly序列中，发现29个精氨酸至少部分甲基化。 Arg-Gly-Gly序列存在于29位甲基化位点中的21个，与其他甲基化蛋白相比，在Gly-Arg-Gly序列中仅发现了11（38％）甲基化精氨酸残基。精氨酸残基的C末端的存在似乎是通过蛋白质 - 精氨酸N-甲基转移酶（PRMT）识别这种不对称二甲基化反应的先决条件。一个单甲基精氨酸和未对称甲基化精氨酸残基。目前的发现表明，RNA结合EWS蛋白的梭子从核以甲基化形式从细胞核到细胞表面，讨论其作用。
In contrast to the knowledge regarding the function of chimeric Ewing sarcoma (EWS) fusion proteins that arise from chromosomal translocation, the cellular function of the RNA binding EWS protein is poorly characterized. EWS protein had been found mainly in the nucleus. In this report we show that EWS protein is not only found in the nucleus and cytosol but also on cell. surfaces. After cell-surface biotinylation, isoelectric focusing of membrane fraction, avidin-agarose extraction of biotinylated proteins, and SDS-polyacrylamide gel electrophoresis, EWS protein was identified by matrix-assisted laser desorption ionization and nanoelectro-spray tandem mass spectrometry of in-gel-digested peptides. These analyses revealed that the protein, having repeated RGG motifs, is extensively asymmetrically dimethylated on arginine residues, the sites of which have been mapped by mass spectrometric methods. Out of a total of 30 Arg-Gly sequences, 29 arginines were found to be at least partially methylated. The Arg-Gly-Gly sequence was present in 21 of the 29 methylation sites, and in contrast to other methylated proteins, only 11 (38%) methylated arginine residues were found in the Gly-Arg-Gly sequence. The presence of Gly on the C-terminal side of the arginine residue seems to be a prerequisite for recognition by a protein-arginine N-methyltransferase (PRMT) catalyzing this asymmetric dimethylation reaction. One monomethylarginine and no symmetrically methylated arginine residue was found. The present findings imply that RNA-binding EWS protein shuttles from the nucleus to the cell surface in a methylated form, the role of which is discussed.