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Chlorotoxin是一个由36个氨基酸组成的线状多肽，可用于氯毒素相关的研究。
Chlorotoxin(linear) is a linear 36 amino-acid peptide which can be used in Chlorotoxin related research.

Definition

Chlorotoxin is a 36-amino acid peptide found in the venom of the death-stalker scorpion (Leiurus quinquestriatus) which blocks small-conductance chloride channels1.

Discovery

Chlorotoxin was first purified from scorpion venum in 19931.

Classification

Chlorotoxin belongs to the scorpion toxin superfamily1.

Structural Characteristics

Chlorotoxin is a small positive charged peptide consisting of 30-35 amino acids and 4 disulfide bonds2. Eight cysteine amino acids among its total number of amino acids present in the structure of this peptide are connected with four disulfide bonds2.

Mode of action

Chlorotoxin specifically binds to matrix-metalloproteinase receptor on glioma cells and inhibits it3. It also acts as a ligand for chloride channels and blocks them4.

Functions

As chlorotoxin binds preferentially to glioma cells compared with non-neoplastic cells or normal brain it has has been used to develop new methods for the treatment and diagnosis of several types of cancer5.  Chlorotoxin: Cy5.5 bioconjugate has been recently used to demarcate cancer cells from the surrounding normal cells. This gives surgeons a better chance of removing all of the cancerous cells without injuring the surrounding healthy tissue6.

References                                      

1.     DeBin JA, Maggio JE, Strichartz GR (1993). Purification and characterization of chlorotoxin, a chloride channel ligand from the venom of the scorpion. Am. J. Physiol., 264 (2 Pt 1), C361–9.

2.     Lippens G, Najib J, Wodak SJ, Tartar A (1995). NMR sequential assignments and solution structure of chlorotoxin, a small scorpion toxin that blocks chloride channels. Biochemistry, 34 (1): 13–21.

3.     Deshane J, Garner CC, Sontheimer H (2003). Chlorotoxin inhibits glioma cell invasion via matrix metalloproteinase-2. J. Biol. Chem., 278 (6), 4135–44.

4.     DeBin JA, Strichartz GR (1991). Chloride channel inhibition by the venom of the scorpion Leiurus quinquestriatue. Toxicon, 29 (11), 1403–8.

5.     Lyons SA, O'Neal J, Sontheimer H (2002). Chlorotoxin, a scorpion-derived peptide, specifically binds to gliomas and tumors of neuroectodermal origin. Glia 39 (2), 162–73.

6.     Veiseh M, Gabikian P, Bahrami SB, et al (2007). Tumor paint: a chlorotoxin: Cy5.5 bioconjugate for intraoperative visualization of cancer foci. Cancer Res., 67 (14), 6882–8.

化学预防肽是有助于预防疾病（例如癌症或糖尿病）的发作或发展的肽。这些肽可以源自天然来源，例如大豆或牛奶，也可以来自肽模拟物的设计，也可以源自使用合成肽进行的肽筛选。据认为，这些肽中的某些可以充当细胞周期的调节剂，其调节使细胞通过复制周期前进所需的蛋白质的产生和功能。另外，现在有越来越多的证据表明特定的饮食模式，食物和饮料以及其他饮食物质可以而且确实可以预防癌症。越来越多的流行病学研究表明，食物，营养和身体活动在预防和改变癌症过程中很重要。包括植物蛋白酶抑制剂，乳铁蛋白，乳铁蛋白，凝集素和lunasin在内的不同类型的食物蛋白和多肽似乎起着化学预防剂的作用。如今，蛋白质和多肽被认为是一组营养保健品，在预防癌症的不同阶段（包括起始，促进和进展）方面显示出潜力。此外，已经发现在植物中发现的一些蛋白酶抑制剂，例如豆类和大豆，是有效的癌发生抑制剂。致癌作用是引发和促进癌症的过程。 Bowman-Birk抑制剂和Kunitz胰蛋白酶抑制剂就在其中。目前，这些化合物在致癌作用中的生物学功能主要归因于抑制癌细胞的侵袭和转移，但是，其作用机理仍不完全清楚，需要进一步研究以充分阐明它们。

多肽H2N-Met-Cys-Met-Pro-Cys-Phe-Thr-Thr-Asp-His-Gln-Met-Ala-Arg-Lys-Cys-Asp-Asp-Cys-Cys-Gly-Gly-Lys-Gly-Arg-Gly-Lys-Cys-Tyr-Gly-Pro-Gln-Cys-Leu-Cys-Arg-NH2的合成步骤：

1、合成MBHA树脂：取若干克的MBHA树脂（如初始取代度为0.5mmol/g）和1倍树脂摩尔量的Fmoc-Linker-OH加入到反应器中，加入DMF，搅拌使氨基酸完全溶解。再加入树脂2倍量的DIEPA，搅拌混合均匀。再加入树脂0.95倍量的HBTU，搅拌混合均匀。反应3-4小时后，用DMF洗涤3次。用2倍树脂体积的10%乙酸酐/DMF 进行封端30分钟。然后再用DMF洗涤3次，甲醇洗涤2次，DCM洗涤2次，再用甲醇洗涤2次。真空干燥12小时以上，得到干燥的树脂{Fmoc-Linker-MHBA Resin}，测定取代度。这里测得取代度为 0.3mmol/g。结构如下图：

2、脱Fmoc：取2.95g的上述树脂，用DCM或DMF溶胀20分钟。用DMF洗涤2遍。加3倍树脂体积的20%Pip/DMF溶液，鼓氮气30分钟，然后2倍树脂体积的DMF 洗涤5次。得到 H2N-Linker-MBHA Resin 。（此步骤脱除Fmoc基团，茚三酮检测为蓝色，Pip为哌啶）。结构图如下：

3、缩合：取2.66mmol Fmoc-Arg(Pbf)-OH 氨基酸，加入到上述树脂里，加适当DMF溶解氨基酸，再依次加入5.31mmol DIPEA，2.52mmol HBTU。反应30分钟后，取小样洗涤，茚三酮检测为无色。用2倍树脂体积的DMF 洗涤3次树脂。（洗涤树脂，去掉残留溶剂，为下一步反应做准备）。得到Fmoc-Arg(Pbf)-Linker-MBHA Resin。氨基酸：DIPEA：HBTU：树脂=3：6：2.85：1（摩尔比）。结构图如下：

4、依次循环步骤二、步骤三，依次得到

H2N-Arg(Pbf)-Linker-MBHA Resin

Fmoc-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

H2N-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

Fmoc-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

H2N-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

Fmoc-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

H2N-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

Fmoc-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

H2N-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

Fmoc-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

H2N-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

Fmoc-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

H2N-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

Fmoc-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

H2N-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

Fmoc-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

H2N-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

Fmoc-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

H2N-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

Fmoc-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

H2N-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

Fmoc-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

H2N-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

Fmoc-Gly-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

H2N-Gly-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

Fmoc-Lys(Boc)-Gly-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

H2N-Lys(Boc)-Gly-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

Fmoc-Gly-Lys(Boc)-Gly-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

H2N-Gly-Lys(Boc)-Gly-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

Fmoc-Gly-Gly-Lys(Boc)-Gly-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

H2N-Gly-Gly-Lys(Boc)-Gly-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

Fmoc-Cys(Trt)-Gly-Gly-Lys(Boc)-Gly-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

H2N-Cys(Trt)-Gly-Gly-Lys(Boc)-Gly-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

Fmoc-Cys(Trt)-Cys(Trt)-Gly-Gly-Lys(Boc)-Gly-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

H2N-Cys(Trt)-Cys(Trt)-Gly-Gly-Lys(Boc)-Gly-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

Fmoc-Asp(OtBu)-Cys(Trt)-Cys(Trt)-Gly-Gly-Lys(Boc)-Gly-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

H2N-Asp(OtBu)-Cys(Trt)-Cys(Trt)-Gly-Gly-Lys(Boc)-Gly-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

Fmoc-Asp(OtBu)-Asp(OtBu)-Cys(Trt)-Cys(Trt)-Gly-Gly-Lys(Boc)-Gly-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

H2N-Asp(OtBu)-Asp(OtBu)-Cys(Trt)-Cys(Trt)-Gly-Gly-Lys(Boc)-Gly-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

Fmoc-Cys(Trt)-Asp(OtBu)-Asp(OtBu)-Cys(Trt)-Cys(Trt)-Gly-Gly-Lys(Boc)-Gly-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

H2N-Cys(Trt)-Asp(OtBu)-Asp(OtBu)-Cys(Trt)-Cys(Trt)-Gly-Gly-Lys(Boc)-Gly-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

Fmoc-Lys(Boc)-Cys(Trt)-Asp(OtBu)-Asp(OtBu)-Cys(Trt)-Cys(Trt)-Gly-Gly-Lys(Boc)-Gly-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

H2N-Lys(Boc)-Cys(Trt)-Asp(OtBu)-Asp(OtBu)-Cys(Trt)-Cys(Trt)-Gly-Gly-Lys(Boc)-Gly-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

Fmoc-Arg(Pbf)-Lys(Boc)-Cys(Trt)-Asp(OtBu)-Asp(OtBu)-Cys(Trt)-Cys(Trt)-Gly-Gly-Lys(Boc)-Gly-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

H2N-Arg(Pbf)-Lys(Boc)-Cys(Trt)-Asp(OtBu)-Asp(OtBu)-Cys(Trt)-Cys(Trt)-Gly-Gly-Lys(Boc)-Gly-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

Fmoc-Ala-Arg(Pbf)-Lys(Boc)-Cys(Trt)-Asp(OtBu)-Asp(OtBu)-Cys(Trt)-Cys(Trt)-Gly-Gly-Lys(Boc)-Gly-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

H2N-Ala-Arg(Pbf)-Lys(Boc)-Cys(Trt)-Asp(OtBu)-Asp(OtBu)-Cys(Trt)-Cys(Trt)-Gly-Gly-Lys(Boc)-Gly-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

Fmoc-Met-Ala-Arg(Pbf)-Lys(Boc)-Cys(Trt)-Asp(OtBu)-Asp(OtBu)-Cys(Trt)-Cys(Trt)-Gly-Gly-Lys(Boc)-Gly-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

H2N-Met-Ala-Arg(Pbf)-Lys(Boc)-Cys(Trt)-Asp(OtBu)-Asp(OtBu)-Cys(Trt)-Cys(Trt)-Gly-Gly-Lys(Boc)-Gly-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

Fmoc-Gln(Trt)-Met-Ala-Arg(Pbf)-Lys(Boc)-Cys(Trt)-Asp(OtBu)-Asp(OtBu)-Cys(Trt)-Cys(Trt)-Gly-Gly-Lys(Boc)-Gly-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

H2N-Gln(Trt)-Met-Ala-Arg(Pbf)-Lys(Boc)-Cys(Trt)-Asp(OtBu)-Asp(OtBu)-Cys(Trt)-Cys(Trt)-Gly-Gly-Lys(Boc)-Gly-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

Fmoc-His(Trt)-Gln(Trt)-Met-Ala-Arg(Pbf)-Lys(Boc)-Cys(Trt)-Asp(OtBu)-Asp(OtBu)-Cys(Trt)-Cys(Trt)-Gly-Gly-Lys(Boc)-Gly-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

H2N-His(Trt)-Gln(Trt)-Met-Ala-Arg(Pbf)-Lys(Boc)-Cys(Trt)-Asp(OtBu)-Asp(OtBu)-Cys(Trt)-Cys(Trt)-Gly-Gly-Lys(Boc)-Gly-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

Fmoc-Asp(OtBu)-His(Trt)-Gln(Trt)-Met-Ala-Arg(Pbf)-Lys(Boc)-Cys(Trt)-Asp(OtBu)-Asp(OtBu)-Cys(Trt)-Cys(Trt)-Gly-Gly-Lys(Boc)-Gly-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

H2N-Asp(OtBu)-His(Trt)-Gln(Trt)-Met-Ala-Arg(Pbf)-Lys(Boc)-Cys(Trt)-Asp(OtBu)-Asp(OtBu)-Cys(Trt)-Cys(Trt)-Gly-Gly-Lys(Boc)-Gly-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

Fmoc-Thr(tBu)-Asp(OtBu)-His(Trt)-Gln(Trt)-Met-Ala-Arg(Pbf)-Lys(Boc)-Cys(Trt)-Asp(OtBu)-Asp(OtBu)-Cys(Trt)-Cys(Trt)-Gly-Gly-Lys(Boc)-Gly-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

H2N-Thr(tBu)-Asp(OtBu)-His(Trt)-Gln(Trt)-Met-Ala-Arg(Pbf)-Lys(Boc)-Cys(Trt)-Asp(OtBu)-Asp(OtBu)-Cys(Trt)-Cys(Trt)-Gly-Gly-Lys(Boc)-Gly-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

Fmoc-Thr(tBu)-Thr(tBu)-Asp(OtBu)-His(Trt)-Gln(Trt)-Met-Ala-Arg(Pbf)-Lys(Boc)-Cys(Trt)-Asp(OtBu)-Asp(OtBu)-Cys(Trt)-Cys(Trt)-Gly-Gly-Lys(Boc)-Gly-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

H2N-Thr(tBu)-Thr(tBu)-Asp(OtBu)-His(Trt)-Gln(Trt)-Met-Ala-Arg(Pbf)-Lys(Boc)-Cys(Trt)-Asp(OtBu)-Asp(OtBu)-Cys(Trt)-Cys(Trt)-Gly-Gly-Lys(Boc)-Gly-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

Fmoc-Phe-Thr(tBu)-Thr(tBu)-Asp(OtBu)-His(Trt)-Gln(Trt)-Met-Ala-Arg(Pbf)-Lys(Boc)-Cys(Trt)-Asp(OtBu)-Asp(OtBu)-Cys(Trt)-Cys(Trt)-Gly-Gly-Lys(Boc)-Gly-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

H2N-Phe-Thr(tBu)-Thr(tBu)-Asp(OtBu)-His(Trt)-Gln(Trt)-Met-Ala-Arg(Pbf)-Lys(Boc)-Cys(Trt)-Asp(OtBu)-Asp(OtBu)-Cys(Trt)-Cys(Trt)-Gly-Gly-Lys(Boc)-Gly-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

Fmoc-Cys(Trt)-Phe-Thr(tBu)-Thr(tBu)-Asp(OtBu)-His(Trt)-Gln(Trt)-Met-Ala-Arg(Pbf)-Lys(Boc)-Cys(Trt)-Asp(OtBu)-Asp(OtBu)-Cys(Trt)-Cys(Trt)-Gly-Gly-Lys(Boc)-Gly-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

H2N-Cys(Trt)-Phe-Thr(tBu)-Thr(tBu)-Asp(OtBu)-His(Trt)-Gln(Trt)-Met-Ala-Arg(Pbf)-Lys(Boc)-Cys(Trt)-Asp(OtBu)-Asp(OtBu)-Cys(Trt)-Cys(Trt)-Gly-Gly-Lys(Boc)-Gly-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

Fmoc-Pro-Cys(Trt)-Phe-Thr(tBu)-Thr(tBu)-Asp(OtBu)-His(Trt)-Gln(Trt)-Met-Ala-Arg(Pbf)-Lys(Boc)-Cys(Trt)-Asp(OtBu)-Asp(OtBu)-Cys(Trt)-Cys(Trt)-Gly-Gly-Lys(Boc)-Gly-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

H2N-Pro-Cys(Trt)-Phe-Thr(tBu)-Thr(tBu)-Asp(OtBu)-His(Trt)-Gln(Trt)-Met-Ala-Arg(Pbf)-Lys(Boc)-Cys(Trt)-Asp(OtBu)-Asp(OtBu)-Cys(Trt)-Cys(Trt)-Gly-Gly-Lys(Boc)-Gly-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

Fmoc-Met-Pro-Cys(Trt)-Phe-Thr(tBu)-Thr(tBu)-Asp(OtBu)-His(Trt)-Gln(Trt)-Met-Ala-Arg(Pbf)-Lys(Boc)-Cys(Trt)-Asp(OtBu)-Asp(OtBu)-Cys(Trt)-Cys(Trt)-Gly-Gly-Lys(Boc)-Gly-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

H2N-Met-Pro-Cys(Trt)-Phe-Thr(tBu)-Thr(tBu)-Asp(OtBu)-His(Trt)-Gln(Trt)-Met-Ala-Arg(Pbf)-Lys(Boc)-Cys(Trt)-Asp(OtBu)-Asp(OtBu)-Cys(Trt)-Cys(Trt)-Gly-Gly-Lys(Boc)-Gly-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

Fmoc-Cys(Trt)-Met-Pro-Cys(Trt)-Phe-Thr(tBu)-Thr(tBu)-Asp(OtBu)-His(Trt)-Gln(Trt)-Met-Ala-Arg(Pbf)-Lys(Boc)-Cys(Trt)-Asp(OtBu)-Asp(OtBu)-Cys(Trt)-Cys(Trt)-Gly-Gly-Lys(Boc)-Gly-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

H2N-Cys(Trt)-Met-Pro-Cys(Trt)-Phe-Thr(tBu)-Thr(tBu)-Asp(OtBu)-His(Trt)-Gln(Trt)-Met-Ala-Arg(Pbf)-Lys(Boc)-Cys(Trt)-Asp(OtBu)-Asp(OtBu)-Cys(Trt)-Cys(Trt)-Gly-Gly-Lys(Boc)-Gly-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

Fmoc-Met-Cys(Trt)-Met-Pro-Cys(Trt)-Phe-Thr(tBu)-Thr(tBu)-Asp(OtBu)-His(Trt)-Gln(Trt)-Met-Ala-Arg(Pbf)-Lys(Boc)-Cys(Trt)-Asp(OtBu)-Asp(OtBu)-Cys(Trt)-Cys(Trt)-Gly-Gly-Lys(Boc)-Gly-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin

以上中间结构，均可在专肽生物多肽计算器-多肽结构计算器中，一键画出。

最后再经过步骤二得到 H2N-Met-Cys(Trt)-Met-Pro-Cys(Trt)-Phe-Thr(tBu)-Thr(tBu)-Asp(OtBu)-His(Trt)-Gln(Trt)-Met-Ala-Arg(Pbf)-Lys(Boc)-Cys(Trt)-Asp(OtBu)-Asp(OtBu)-Cys(Trt)-Cys(Trt)-Gly-Gly-Lys(Boc)-Gly-Arg(Pbf)-Gly-Lys(Boc)-Cys(Trt)-Tyr(tBu)-Gly-Pro-Gln(Trt)-Cys(Trt)-Leu-Cys(Trt)-Arg(Pbf)-Linker-MBHA Resin，结构如下：

5、切割：6倍树脂体积的切割液（或每1g树脂加8ml左右的切割液），摇床摇晃 2小时，过滤掉树脂，用冰无水乙醚沉淀滤液，并用冰无水乙醚洗涤沉淀物3次，最后将沉淀物放真空干燥釜中，常温干燥24小试，得到粗品H2N-Met-Cys-Met-Pro-Cys-Phe-Thr-Thr-Asp-His-Gln-Met-Ala-Arg-Lys-Cys-Asp-Asp-Cys-Cys-Gly-Gly-Lys-Gly-Arg-Gly-Lys-Cys-Tyr-Gly-Pro-Gln-Cys-Leu-Cys-Arg-NH2。结构图见产品结构图。

切割液选择：1）TFA：H2O=95%：5%

2）TFA：H2O：TIS=95%：2.5%：2.5%

3）三氟乙酸：茴香硫醚：1，2-乙二硫醇：苯酚：水=87.5%：5%：2.5%：2.5%：2.5%

（前两种适合没有容易氧化的氨基酸，例如Trp、Cys、Met。第三种适合几乎所有的序列。）

6、纯化冻干：使用液相色谱纯化，收集目标峰液体，进行冻干，获得蓬松的粉末状固体多肽。不过这时要取小样复测下纯度 是否目标纯度。

7、最后总结：

杭州专肽生物技术有限公司（ALLPEPTIDE https://www.allpeptide.com）主营定制多肽合成业务，提供各类长肽，短肽，环肽，提供各类修饰肽，如：荧光标记修饰（CY3、CY5、CY5.5、CY7、FAM、FITC、Rhodamine B、TAMRA等），功能基团修饰肽（叠氮、炔基、DBCO、DOTA、NOTA等），同位素标记肽（N15、C13），订书肽（Stapled Peptide）,脂肪酸修饰肽（Pal、Myr、Ste），磷酸化修饰肽（P-Ser、P-Thr、P-Tyr），环肽（酰胺键环肽、一对或者多对二硫键环），生物素标记肽，PEG修饰肽，甲基化修饰肽等。

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