Peptide backbone conformation affects the substrate preference of protein arginine methyltransferase I.
Peptide backbone conformation affects the substrate preference of protein arginine methyltransferase I.
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DOI：
10.1021/bi300373b
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发表时间：
2012-06
期刊：
影响因子：
2.9
通讯作者：
Knut Kölbel;C. Ihling;U. Kühn;I. Neundorf;S. Otto;Jan Stichel;D. Robaa;A. Beck‐Sickinger;A. Sinz;E. Wahle
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文献类型：
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作者：
Knut Kölbel;C. Ihling;U. Kühn;I. Neundorf;S. Otto;Jan Stichel;D. Robaa;A. Beck‐Sickinger;A. Sinz;E. Wahle
精氨酸侧链的不对称二甲基化是真核生物蛋白质的一种常见的翻译后修饰，主要用于调节蛋白质之间的相互作用。这种修饰是由I型蛋白精氨酸甲基转移酶催化的，PRMT1是该家族的主要成员。这些酶的底物专一性的决定因素还知之甚少。核多聚(A)结合蛋白1(PABPN1)是由位于蛋白质C-末端结构域RXR序列中13个精氨酸残基的PRMT1甲基化而成。我们已经在PABPN1和相应的合成肽中确定了PRMT1催化甲基化的首选位置。用稳态动力学分析和质谱仪分析了这些底物的变异体。数据表明，最初的甲基化是由N-末端相邻的脯氨酸指向首选的精氨酸残基。多肽环化时的甲基化增强表明，反向转折结构的诱导是相应的脯氨酸残基能够优先甲基化相邻精氨酸残基的基础，这一观点得到了远紫外圆二色谱的支持。我们认为，反转的形成有利于精氨酸侧链进入PRMT1的活性部位，这些活性部位位于甜甜圈状PRMT1同源二聚体的中心空腔中。
Asymmetric dimethylation of arginine side chains is a common post-translational modification of eukaryotic proteins, which serves mostly to regulate protein-protein interactions. The modification is catalyzed by type I protein arginine methyltransferases, PRMT1 being the predominant member of the family. Determinants of substrate specificity of these enzymes are poorly understood. The Nuclear poly(A) binding protein 1 (PABPN1) is methylated by PRMT1 at 13 arginine residues located in RXR sequences in the protein's C-terminal domain. We have identified a preferred site for PRMT1-catalyzed methylation in PABPN1 and in a corresponding synthetic peptide. Variants of these substrates were analyzed by steady-state kinetic analysis and mass spectrometry. The data indicate that initial methylation is directed toward the preferred arginine residue by an N-terminally adjacent proline. Enhanced methylation upon peptide cyclization suggests that induction of a reverse turn structure is the basis for the ability of the respective proline residue to enable preferred methylation of the neighboring arginine residue, and this notion is supported by far-UV circular dichroism spectroscopy. We suggest that the formation of a reverse turn facilitates the access of arginine side chains to the active sites of PRMT1, which are located in the central cavity of a doughnut-shaped PRMT1 homodimer.