400-998-5282
专注多肽 服务科研
400-998-5282
专注多肽 服务科研
淀粉样蛋白 Bri 蛋白 (1-34) 由位于 13 号染色体上的 BRI 基因编码，以淀粉样原纤维形式沉积，导致神经元功能障碍和痴呆。
编号：200041
CAS号：321853-88-7
单字母：Pyr-ASNCFAIRHFENKFAVETLICSRTVKKNIIEEN-OH
Amyloid Bri Protein (1-34), encoded by the gene BRI located on chromosome 13, is deposited as amyloid fibrils causing neuronal dysfunction and dementia.
| DOI | 名称 | |
|---|---|---|
| 10.1038/21637 | A stop-codon mutation in the BRI gene associated with familial British dementia | 下载 |
多肽Pyr-Ala-Ser-Asn-Cys-Phe-Ala-Ile-Arg-His-Phe-Glu-Asn-Lys-Phe-Ala-Val-Glu-Thr-Leu-Ile-Cys-Ser-Arg-Thr-Val-Lys-Lys-Asn-Ile-Ile-Glu-Glu-Asn-COOH的合成步骤：
1、合成CTC树脂：称取2.76g CTC Resin（如初始取代度约为1.2mmol/g）和3.97mmol Fmoc-Asn(Trt)-OH于反应器中，加入适量DCM溶解氨基酸（需要注意，此时CTC树脂体积会增大好几倍，避免DCM溶液过少），再加入9.94mmol DIPEA（Mw：129.1,d：0.740g/ml），反应2-3小时后，可不抽滤溶液，直接加入1ml的HPLC级甲醇，封端半小时。依次用DMF洗涤2次，甲醇洗涤1次，DCM洗涤一次，甲醇洗涤一次，DCM洗涤一次，DMF洗涤2次（这里使用甲醇和DCM交替洗涤，是为了更好地去除其他溶质，有利于后续反应）。得到 Fmoc-Asn(Trt)-CTC Resin。结构图如下：
2、脱Fmoc：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-Asn(Trt)-CTC 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9.94mmol Fmoc-Glu(OtBu)-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19.87mmol DIPEA，9.44mmol 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-Glu(OtBu)-Asn(Trt)-CTC 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-Glu(OtBu)-Asn(Trt)-CTC Resin
Fmoc-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
H2N-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
Fmoc-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
H2N-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
Fmoc-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
H2N-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
Fmoc-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
H2N-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
Fmoc-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
H2N-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
Fmoc-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
H2N-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
Fmoc-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
H2N-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
Fmoc-Thr(tBu)-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
H2N-Thr(tBu)-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
Fmoc-Arg(Pbf)-Thr(tBu)-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
H2N-Arg(Pbf)-Thr(tBu)-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
Fmoc-Ser(tBu)-Arg(Pbf)-Thr(tBu)-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
H2N-Ser(tBu)-Arg(Pbf)-Thr(tBu)-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
Fmoc-Cys(Trt)-Ser(tBu)-Arg(Pbf)-Thr(tBu)-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
H2N-Cys(Trt)-Ser(tBu)-Arg(Pbf)-Thr(tBu)-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
Fmoc-Ile-Cys(Trt)-Ser(tBu)-Arg(Pbf)-Thr(tBu)-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
H2N-Ile-Cys(Trt)-Ser(tBu)-Arg(Pbf)-Thr(tBu)-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
Fmoc-Leu-Ile-Cys(Trt)-Ser(tBu)-Arg(Pbf)-Thr(tBu)-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
H2N-Leu-Ile-Cys(Trt)-Ser(tBu)-Arg(Pbf)-Thr(tBu)-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
Fmoc-Thr(tBu)-Leu-Ile-Cys(Trt)-Ser(tBu)-Arg(Pbf)-Thr(tBu)-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
H2N-Thr(tBu)-Leu-Ile-Cys(Trt)-Ser(tBu)-Arg(Pbf)-Thr(tBu)-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
Fmoc-Glu(OtBu)-Thr(tBu)-Leu-Ile-Cys(Trt)-Ser(tBu)-Arg(Pbf)-Thr(tBu)-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
H2N-Glu(OtBu)-Thr(tBu)-Leu-Ile-Cys(Trt)-Ser(tBu)-Arg(Pbf)-Thr(tBu)-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
Fmoc-Val-Glu(OtBu)-Thr(tBu)-Leu-Ile-Cys(Trt)-Ser(tBu)-Arg(Pbf)-Thr(tBu)-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
H2N-Val-Glu(OtBu)-Thr(tBu)-Leu-Ile-Cys(Trt)-Ser(tBu)-Arg(Pbf)-Thr(tBu)-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
Fmoc-Ala-Val-Glu(OtBu)-Thr(tBu)-Leu-Ile-Cys(Trt)-Ser(tBu)-Arg(Pbf)-Thr(tBu)-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
H2N-Ala-Val-Glu(OtBu)-Thr(tBu)-Leu-Ile-Cys(Trt)-Ser(tBu)-Arg(Pbf)-Thr(tBu)-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
Fmoc-Phe-Ala-Val-Glu(OtBu)-Thr(tBu)-Leu-Ile-Cys(Trt)-Ser(tBu)-Arg(Pbf)-Thr(tBu)-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
H2N-Phe-Ala-Val-Glu(OtBu)-Thr(tBu)-Leu-Ile-Cys(Trt)-Ser(tBu)-Arg(Pbf)-Thr(tBu)-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
Fmoc-Lys(Boc)-Phe-Ala-Val-Glu(OtBu)-Thr(tBu)-Leu-Ile-Cys(Trt)-Ser(tBu)-Arg(Pbf)-Thr(tBu)-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
H2N-Lys(Boc)-Phe-Ala-Val-Glu(OtBu)-Thr(tBu)-Leu-Ile-Cys(Trt)-Ser(tBu)-Arg(Pbf)-Thr(tBu)-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
Fmoc-Asn(Trt)-Lys(Boc)-Phe-Ala-Val-Glu(OtBu)-Thr(tBu)-Leu-Ile-Cys(Trt)-Ser(tBu)-Arg(Pbf)-Thr(tBu)-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
H2N-Asn(Trt)-Lys(Boc)-Phe-Ala-Val-Glu(OtBu)-Thr(tBu)-Leu-Ile-Cys(Trt)-Ser(tBu)-Arg(Pbf)-Thr(tBu)-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
Fmoc-Glu(OtBu)-Asn(Trt)-Lys(Boc)-Phe-Ala-Val-Glu(OtBu)-Thr(tBu)-Leu-Ile-Cys(Trt)-Ser(tBu)-Arg(Pbf)-Thr(tBu)-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
H2N-Glu(OtBu)-Asn(Trt)-Lys(Boc)-Phe-Ala-Val-Glu(OtBu)-Thr(tBu)-Leu-Ile-Cys(Trt)-Ser(tBu)-Arg(Pbf)-Thr(tBu)-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
Fmoc-Phe-Glu(OtBu)-Asn(Trt)-Lys(Boc)-Phe-Ala-Val-Glu(OtBu)-Thr(tBu)-Leu-Ile-Cys(Trt)-Ser(tBu)-Arg(Pbf)-Thr(tBu)-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
H2N-Phe-Glu(OtBu)-Asn(Trt)-Lys(Boc)-Phe-Ala-Val-Glu(OtBu)-Thr(tBu)-Leu-Ile-Cys(Trt)-Ser(tBu)-Arg(Pbf)-Thr(tBu)-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
Fmoc-His(Trt)-Phe-Glu(OtBu)-Asn(Trt)-Lys(Boc)-Phe-Ala-Val-Glu(OtBu)-Thr(tBu)-Leu-Ile-Cys(Trt)-Ser(tBu)-Arg(Pbf)-Thr(tBu)-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
H2N-His(Trt)-Phe-Glu(OtBu)-Asn(Trt)-Lys(Boc)-Phe-Ala-Val-Glu(OtBu)-Thr(tBu)-Leu-Ile-Cys(Trt)-Ser(tBu)-Arg(Pbf)-Thr(tBu)-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
Fmoc-Arg(Pbf)-His(Trt)-Phe-Glu(OtBu)-Asn(Trt)-Lys(Boc)-Phe-Ala-Val-Glu(OtBu)-Thr(tBu)-Leu-Ile-Cys(Trt)-Ser(tBu)-Arg(Pbf)-Thr(tBu)-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
H2N-Arg(Pbf)-His(Trt)-Phe-Glu(OtBu)-Asn(Trt)-Lys(Boc)-Phe-Ala-Val-Glu(OtBu)-Thr(tBu)-Leu-Ile-Cys(Trt)-Ser(tBu)-Arg(Pbf)-Thr(tBu)-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
Fmoc-Ile-Arg(Pbf)-His(Trt)-Phe-Glu(OtBu)-Asn(Trt)-Lys(Boc)-Phe-Ala-Val-Glu(OtBu)-Thr(tBu)-Leu-Ile-Cys(Trt)-Ser(tBu)-Arg(Pbf)-Thr(tBu)-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
H2N-Ile-Arg(Pbf)-His(Trt)-Phe-Glu(OtBu)-Asn(Trt)-Lys(Boc)-Phe-Ala-Val-Glu(OtBu)-Thr(tBu)-Leu-Ile-Cys(Trt)-Ser(tBu)-Arg(Pbf)-Thr(tBu)-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
Fmoc-Ala-Ile-Arg(Pbf)-His(Trt)-Phe-Glu(OtBu)-Asn(Trt)-Lys(Boc)-Phe-Ala-Val-Glu(OtBu)-Thr(tBu)-Leu-Ile-Cys(Trt)-Ser(tBu)-Arg(Pbf)-Thr(tBu)-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
H2N-Ala-Ile-Arg(Pbf)-His(Trt)-Phe-Glu(OtBu)-Asn(Trt)-Lys(Boc)-Phe-Ala-Val-Glu(OtBu)-Thr(tBu)-Leu-Ile-Cys(Trt)-Ser(tBu)-Arg(Pbf)-Thr(tBu)-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
Fmoc-Phe-Ala-Ile-Arg(Pbf)-His(Trt)-Phe-Glu(OtBu)-Asn(Trt)-Lys(Boc)-Phe-Ala-Val-Glu(OtBu)-Thr(tBu)-Leu-Ile-Cys(Trt)-Ser(tBu)-Arg(Pbf)-Thr(tBu)-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
H2N-Phe-Ala-Ile-Arg(Pbf)-His(Trt)-Phe-Glu(OtBu)-Asn(Trt)-Lys(Boc)-Phe-Ala-Val-Glu(OtBu)-Thr(tBu)-Leu-Ile-Cys(Trt)-Ser(tBu)-Arg(Pbf)-Thr(tBu)-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
Fmoc-Cys(Trt)-Phe-Ala-Ile-Arg(Pbf)-His(Trt)-Phe-Glu(OtBu)-Asn(Trt)-Lys(Boc)-Phe-Ala-Val-Glu(OtBu)-Thr(tBu)-Leu-Ile-Cys(Trt)-Ser(tBu)-Arg(Pbf)-Thr(tBu)-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
H2N-Cys(Trt)-Phe-Ala-Ile-Arg(Pbf)-His(Trt)-Phe-Glu(OtBu)-Asn(Trt)-Lys(Boc)-Phe-Ala-Val-Glu(OtBu)-Thr(tBu)-Leu-Ile-Cys(Trt)-Ser(tBu)-Arg(Pbf)-Thr(tBu)-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
Fmoc-Asn(Trt)-Cys(Trt)-Phe-Ala-Ile-Arg(Pbf)-His(Trt)-Phe-Glu(OtBu)-Asn(Trt)-Lys(Boc)-Phe-Ala-Val-Glu(OtBu)-Thr(tBu)-Leu-Ile-Cys(Trt)-Ser(tBu)-Arg(Pbf)-Thr(tBu)-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
H2N-Asn(Trt)-Cys(Trt)-Phe-Ala-Ile-Arg(Pbf)-His(Trt)-Phe-Glu(OtBu)-Asn(Trt)-Lys(Boc)-Phe-Ala-Val-Glu(OtBu)-Thr(tBu)-Leu-Ile-Cys(Trt)-Ser(tBu)-Arg(Pbf)-Thr(tBu)-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
Fmoc-Ser(tBu)-Asn(Trt)-Cys(Trt)-Phe-Ala-Ile-Arg(Pbf)-His(Trt)-Phe-Glu(OtBu)-Asn(Trt)-Lys(Boc)-Phe-Ala-Val-Glu(OtBu)-Thr(tBu)-Leu-Ile-Cys(Trt)-Ser(tBu)-Arg(Pbf)-Thr(tBu)-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
H2N-Ser(tBu)-Asn(Trt)-Cys(Trt)-Phe-Ala-Ile-Arg(Pbf)-His(Trt)-Phe-Glu(OtBu)-Asn(Trt)-Lys(Boc)-Phe-Ala-Val-Glu(OtBu)-Thr(tBu)-Leu-Ile-Cys(Trt)-Ser(tBu)-Arg(Pbf)-Thr(tBu)-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin
Fmoc-Ala-Ser(tBu)-Asn(Trt)-Cys(Trt)-Phe-Ala-Ile-Arg(Pbf)-His(Trt)-Phe-Glu(OtBu)-Asn(Trt)-Lys(Boc)-Phe-Ala-Val-Glu(OtBu)-Thr(tBu)-Leu-Ile-Cys(Trt)-Ser(tBu)-Arg(Pbf)-Thr(tBu)-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC 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-Ala-Ser(tBu)-Asn(Trt)-Cys(Trt)-Phe-Ala-Ile-Arg(Pbf)-His(Trt)-Phe-Glu(OtBu)-Asn(Trt)-Lys(Boc)-Phe-Ala-Val-Glu(OtBu)-Thr(tBu)-Leu-Ile-Cys(Trt)-Ser(tBu)-Arg(Pbf)-Thr(tBu)-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTC Resin，结构如下：
5、焦谷氨酸反应连接：在上述树脂中，加入适当DMF后，再加入9.94mmol 焦谷氨酸到树脂中，再加入19.87mmol DIPEA、9.44mmol HBTU，鼓氮气反应30分钟。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Pyr-Ala-Ser(tBu)-Asn(Trt)-Cys(Trt)-Phe-Ala-Ile-Arg(Pbf)-His(Trt)-Phe-Glu(OtBu)-Asn(Trt)-Lys(Boc)-Phe-Ala-Val-Glu(OtBu)-Thr(tBu)-Leu-Ile-Cys(Trt)-Ser(tBu)-Arg(Pbf)-Thr(tBu)-Val-Lys(Boc)-Lys(Boc)-Asn(Trt)-Ile-Ile-Glu(OtBu)-Glu(OtBu)-Asn(Trt)-CTCResin。 结构如下：
6、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Pyr-Ala-Ser-Asn-Cys-Phe-Ala-Ile-Arg-His-Phe-Glu-Asn-Lys-Phe-Ala-Val-Glu-Thr-Leu-Ile-Cys-Ser-Arg-Thr-Val-Lys-Lys-Asn-Ile-Ile-Glu-Glu-Asn-COOH。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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