Development of Practical Synthetic Method for Phosphopeptides using Two-step Deprotecting Procedures

已经开发了一种方案，用于合成含有o -磷酸化酪氨酸、丝氨酸和/或苏氨酸的肽。该过程包括使用Boc化学将二甲基保护的磷酸化氨基酸衍生物纳入肽中，随后使用两步脱保护方法去除Me基团，包括高酸性和低酸性处理。建立了保护树脂的最佳脱保护条件，即第一步试剂（TFA、间甲酚、EDT中的1 M tmsotf -硫代苯甲醚）和第二步试剂（第一步试剂+ DMS-TMSOTf）的组合。此外，第二步试剂可以改为更有效的试剂体系（tmsotf - dms -m-甲酚- edt）。该合成方法产率高，且无明显副反应。为了合成磷酸化蛋白，我们开发了sn1脱保护程序（tfmsa - tfa -m-甲酚）。用该体系对受保护的磷酸肽硫酯树脂进行处理，得到受保护的磷酸肽硫酯，其中Lys侧链和磷酸氨基侧链分别被CLZ或Me基团保护。利用保护性磷酸肽硫酯合成磷蛋白的研究正在进行中。

A protocol has been developed for the synthesis of peptides containing O-phosphorylated tyrosines, serines, and/or threonines. The procedure involves incorporation of dimethyl-protected phosphorylated amino acid derivatives into peptides using Boc chemistry and subsequent removal of Me groups using a two-step deprotection method consisting of high acidic and low acidic treatments. Optimized deprotection conditions for the protected resins were established, which consist of a combination of the first-step reagent (1 M TMSOTf-thioanisole in TFA,m-cresol, EDT) and the second-step reagent (first-step reagent + DMS-TMSOTf). Furthermore, the second-step reagent can be changed to the more effective reagent system (TMSOTf-DMS-m-cresol-EDT). Theses synthetic method afforded phosphoamino acid-containing peptide in high yield without significant accompanying side reactions. For a phosphoprotein synthesis, we developed SN1-deprotecting procedure (TFMSA-TFA-m-cresol). Treatment of protcted phosphopeptide thioester resins with this system afforded protected phosphopeptide thioesters, in which side chains of Lys and phosphoamino were protected with CLZ or Me grpoups, respectively. Synthetic studies on phosphoprotein utilizing protected phosphopeptide thioesters are in progress.