有机化学 ›› 2017, Vol. 37 ›› Issue (9): 2409-2415.DOI: 10.6023/cjoc201704020 上一篇    下一篇

研究简报

半选择性氧化形成三对二硫键合成利那洛肽

葛巍巍a,b, 陈静a, 张也a, 宗良a, 张鸣a, 董俊军a   

  1. a 中国人民解放军防化学院生化防护系 北京 102205;
    b 公安消防部队高等专科学校训练部 昆明 650208
  • 收稿日期:2017-04-12 修回日期:2017-05-15 出版日期:2017-09-25 发布日期:2017-05-25
  • 通讯作者: 董俊军 E-mail:dongjj536@sina.com
  • 基金资助:

    毒药物与毒理学国家重点实验室开放基金(No.PMC201507)和国民核生化灾害防护国家重点实验室开放基金(No.SKLNBC2013-01K)资助项目.

Semiregioselective Formation of Linaclotide with Orthogonal Cysteine Protection Strategy

Ge Weiweia,b, Chen Jinga, Zhang Yea, Zong Lianga, Zhang Minga, Dong Junjuna   

  1. a Department of Bio-Chem Defence, Institute of Nuclear, Biological and Chemical Defence, Beijing 102205;
    b Department of Training, Public Security Fire Forces College, Kunming 650208
  • Received:2017-04-12 Revised:2017-05-15 Online:2017-09-25 Published:2017-05-25
  • Contact: 10.6023/cjoc201704020 E-mail:dongjj536@sina.com
  • Supported by:

    Project supported by the State Key Laboratory of Toxicology and Medical Countermeasures of China (No. PMC201507) and the State Key Laboratory of Nuclear, Biological and Chemical Protection for Civilian (No. SKLNBC2013-01K).

利用Fmoc固相合成策略,Wang树脂为载体,使用三苯甲基(Trt)和乙酰胺甲基(Acm)保护基的半胱氨酸合成了3条[4 Trt+2 Acm]和3条[2 Trt+4 Acm]利那洛肽的线性前体化合物.在此基础上,采用半选择性氧化策略合成含有三对二硫键的利那洛肽.首先使用含三氟乙酸(TFA)的裂解剂脱除线性前体肽中半胱氨酸的Trt保护基,并使用氯化血红素催化氧化半胱氨酸自由巯基形成二硫键.下一步使用PhS(O) Ph/CH3SiCl3试剂体系脱除剩余保护半胱氨酸的Acm保护基,并同时形成二硫键.使用这种策略,在6条线性前体肽中,有3条可以得到利那洛肽,转化率分别为71.9%、31.5%、81.4%.通过分析6条线性前体肽中二硫键形成的先后顺序对目标产物生成的影响,发现二硫键Cys5-Cys13的形成对利那洛肽的氧化折叠非常关键,在选择性氧化合成利那洛肽时应当优先形成这对二硫键.

关键词: 利那洛肽, 二硫键, 氧化, 半选择性

Six linear precursors of linaclotide containing different protected cysteine residues were synthesized by Fmoc solid-phase methods. Wang resin was used in the peptide syntheses. The protective groups of cysteine thiol were trityl (Trt) and acetamidomethyl (Acm) in the different positions. The six linear precursors of linaclotide include three[4 Trt+2 Acm] and three[2 Trt+4 Acm] ones. The linaclotide with three disulfide bonds was prepared from these linear precursors by semiregioselective strategy. Firstly, linear peptides were cleaved from Wang resins by TFA-TIS-H2O. At the same time, the Trt groups were removed to give free thiol groups, whereas Acm groups were still remained in the peptides. Secondly, the free thiol groups were oxidized by 20% hemin/DIEA system to from disulfide bond(s). Finally, cysteines containing Acm groups were deprotected by CH3SiCl3/PhS(O)Ph/TFA coaktail and disulfide bond(s) were formed simultaneously. The precursors of peptides[4 Trt(2,5,10,13)+2 Acm(1,6)],[2 Trt(1,6)+4 Acm(2,5,10,13)] and[2 Trt(5,13)+2 Acm(1,2,6,10)] give linaclotide at the conversion ratios of 71.9%, 31.5%, and 81.4% respectively. Other three peptides failed in the conversion or were found to be less suitable to prepare linaclotide. Our results indicated that the order of disulfide bond formation is very important to prepare linaclotide by using semiregioselective or regioselective strategies. The Cys5-Cys13 disulfide bond is the most privileged and should be formed firstly among the three disulfide bonds in linaclotide.

Key words: linaclotide, disulfide, oxidation, semiregioselective