Identification of highly methylated arginine residues in an endogenous 20-kDa polypeptide in cancer cells
Identification of highly methylated arginine residues in an endogenous 20-kDa polypeptide in cancer cells
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DOI：
10.1016/s0024-3205(99)00300-8
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发表时间：
1999-07-16
期刊：
影响因子：
6.1
通讯作者：
Kim, S
中科院分区：
文献类型：
--
作者：
Gu, HM;Park, SH;Kim, S
研究了几种癌细胞系中内源性蛋白质的酶促甲基化，以了解蛋白质-精氨酸甲基化与细胞增殖之间的可能关系。从几种癌细胞（HeLa、HCT-48、A549和HepG 2）制备的胞质提取物与S-腺苷-L-[甲基-H-3]甲硫氨酸孵育后，显示出强烈的[甲基-H-3]-标记的20-kDa多肽。另一方面，从正常结肠细胞制备的胞质提取物在相同条件下未显示任何20 kDa蛋白的甲基化。为了鉴定20-kDa多肽的性质，用HCT-48胞质提取物将纯化的组蛋白甲基化，并通过SDS-PAGE进行分析。然而，没有一个组蛋白与甲基化的20-kDa多肽共迁移，表明它不太可能是任何组蛋白亚类。20-kDa多肽中的[甲基-H-3]基团在pH 10-11（37 ℃，30分钟）下稳定，并且甲基化不受GTP γ S（4 mM）刺激，因此反应既不是异戊酰基残基上的羧基甲基化，也不是C-末端法尼基化半胱氨酸上的羧基甲基化。本研究连同先前在HCT-48胞质溶胶组分中鉴定的N-G-甲基化精氨酸残基表明，这种新型内源性20-kDa精氨酸甲基化是细胞增殖相关的翻译后修饰反应。
Enzymatic methylation of endogenous proteins in several cancer cell lines was investigated to understand a possible relationship between protein-arginine methylation and cellular proliferation. Cytosolic extracts prepared from several cancer cells (HeLa, HCT-48, A549, and HepG2) and incubated with S-adenosyl-L-[methyl-H-3]methionine revealed an intensely [methyl-H-3]-labeled 20-kDa polypeptide. On the other hand, cytosolic extracts prepared from normal colon cells did not show any methylation of the 20-kDa protein under identical conditions. To identify nature of the 20-kDa polypeptide, purified histones were methylated with HCT-48 cytosolic extracts and analyzed by SDS-PAGE. However, none of the histones comigrated with the methylated 20-kDa polypeptide, indicating that it is unlikely to be any of the histone subclasses. The [methyl-H-3]group in the 20-kDa polypeptide was stable at pH 10-11 (37 degrees C for 30 min) and methylation was not stimulated by GTP gamma S (4 mM), thus the reaction is neither carboxyl methylesterification on isoaspartyl residues, nor on C-terminal farnesylated cysteine. The present study together with the previous identification of N-G-methylated arginine residues in the HCT-48 cytosol fraction suggests that this novel endogenous 20-kDa arginine-methylation is a cellular proliferation-related posttranslational modification reaction.