Probing Peptidylprolyl Bond cis/trans Status Using Distal 19 F NMR Reporters.
Probing Peptidylprolyl Bond cis/trans Status Using Distal 19 F NMR Reporters.
复制标题
使用远端 19 F NMR 报告仪探测肽基脯氨酰键顺式/反式状态。
DOI：
10.1002/chem.202203017
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发表时间：
2023
期刊：
影响因子：
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通讯作者：
Killoran PM
中科院分区：
文献类型：
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作者：
Killoran PM
本文描述了一种用4-氟苯丙氨酸(4FP)作为~(19)F核磁共振远端报告分子的方法来测量多肽模型中的肽基-脯氨酸键的构象状态。在pH 7.4时，测定了总结构为Ac-X-Pro-Z-Ala-Ala-4FPhe(X和Z为蛋白生成氨基酸)的多肽的顺式-Pro基团百分比，提供的构象基团与用更复杂的方法获得的文献值一致。该方法被应用于探索α-突触核蛋白本质无序的C-末端区域的五肽模型中的Pro键状态，该模型反映了Ac-X-Pro-Z-Ala-4FPhe模型中的偏好。有利的是，19F报告基团不需要邻近或连接到Pro来提供可量化的信号，并且可以放置远端的4-氟苯丙氨酸以不影响Pro键的构象。最后，我们证明了当脯氨酸的羧基上有可电离的氨基酸残基时，脯氨酸键的状态不受pH的显著影响，这使得~(19)F核磁共振成为研究不同pH范围以及不同溶剂和缓冲液中的脯氨酸异构化的宝贵工具。
A method for measuring peptidylprolyl bondcis‐transconformational status in peptide models is described, using 4‐fluorophenylalanine (4FPhe) as a distal reporter for19F NMR. The %cis‐Pro population was measured for peptides of the general structure Ac‐X‐Pro‐Z‐Ala‐Ala‐4FPhe (X and Z are proteinogenic amino acids) at pH 7.4, and provided conformational populations consistent with literature values obtained by more complex methods. This approach was applied to probe the prolyl bond status in pentapeptide models of the intrinsically disordered C‐terminal region of α‐synuclein, which mirrored the preferences in the Ac‐X‐Pro‐Z‐Ala‐4FPhe models. Advantageously, the19F reporter group does not need to be adjacent to or attached to proline to provide quantifiable signals and distal 4‐fluorophenylalanines can be placed so as not to influence prolyl bond conformation. Finally, we demonstrated that the prolyl bond status is not significantly affected by pH when there are ionisable amino acid residues at the carboxyl side of proline, which makes19F NMR an invaluable tool with which to study proline isomerism at a range of pHs and in different solvents and buffers.