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ASV-30 是一種構(gòu)象受限的 sauvagine 類似物,是一種針對小鼠 CRF 受體 2β (mCRFR2β) 的高效選擇性拮抗劑。它已被證明可抑制表達(dá) mCRFR2β 的人胚胎腎 (HEK) 293 細(xì)胞中激動劑誘導(dǎo)的腺苷酸環(huán)化酶活性。
編號:136970
CAS號:220673-95-0
單字母:H2N-fHLLRKMIEIEKQEKEKQQAANNRLLLDTI-CONH2
編號: | 136970 |
中文名稱: | Antisauvagine-30 |
英文名: | Antisauvagine-30 |
英文同義詞: | (D-Phe11,His12)-Sauvagine (11-40);11-D-Phenylalanine-12-L-histidine-11-40-sauvagine (11-40) |
CAS號: | 220673-95-0 |
單字母: | H2N-fHLLRKMIEIEKQEKEKQQAANNRLLLDTI-CONH2 |
三字母: | H2N N端氨基 -DPheD型笨丙氨酸 -His組氨酸 -Leu亮氨酸 -Leu亮氨酸 -Arg精氨酸 -Lys賴氨酸 -Met甲硫氨酸 -Ile異亮氨酸 -Glu谷氨酸 -Ile異亮氨酸 -Glu谷氨酸 -Lys賴氨酸 -Gln谷氨酰胺 -Glu谷氨酸 -Lys賴氨酸 -Glu谷氨酸 -Lys賴氨酸 -Gln谷氨酰胺 -Gln谷氨酰胺 -Ala丙氨酸 -Ala丙氨酸 -Asn天冬酰胺 -Asn天冬酰胺 -Arg精氨酸 -Leu亮氨酸 -Leu亮氨酸 -Leu亮氨酸 -Asp天冬氨酸 -Thr蘇氨酸 -Ile異亮氨酸 -CONH2C端酰胺化 |
氨基酸個數(shù): | 30 |
分子式: | C161H274N48O46S1 |
平均分子量: | 3650.26 |
精確分子量: | 3648.03 |
等電點(PI): | 10.33 |
pH=7.0時的凈電荷數(shù): | 2.22 |
平均親水性: | 0.46333333333333 |
疏水性值: | -0.76 |
外觀與性狀: | 白色粉末狀固體 |
消光系數(shù): | - |
來源: | 人工化學(xué)合成,僅限科學(xué)研究使用,不得用于人體。 |
純度: | 95%、98% |
鹽體系: | 可選TFA、HAc、HCl |
儲存條件: | 負(fù)80℃至負(fù)20℃ |
標(biāo)簽: | D型氨基酸肽 |
ASV-30, a conformationally constrained analog of sauvagine, is a highly potent and selective antagonist directed against mouse CRF receptor type 2β (mCRFR2β). It has been shown to suppress agonist-induced adenylate cyclase activity in human embryonic kidney (HEK) 293 cells expressing mCRFR2β.
Antisauvagine-30是一種選擇性、競爭性和強效的促腎上腺皮質(zhì)激素釋放因子CRF2受體拮抗劑,與大鼠CRF1和小鼠CRF2β受體結(jié)合的Kd值分別為153.6和1.4nM。它可以防止小鼠體內(nèi)應(yīng)激增強的恐懼條件和ERK1/2的MEK1/2依賴性激活。它抑制陰道刺激的cAMP在HEK-mCRF2β細(xì)胞中的積累。
Antisauvagine-30 is a selective, competitive and potent corticotropin-releasing factor CRF2 receptor antagonist with Kd values of 153.6 and 1.4 nM for binding to rat CRF1 and mouse CRF2β receptors respectively. It prevents stress-enhanced fear conditioning and MEK 1/2-dependent activation of ERK1/2 in mice in vivo. It inhibits sauvagine-stimulated cAMP accumulation in HEK-mCRF2β cells.
很多蛋白在細(xì)胞中非常容易被降解,或被標(biāo)記,進(jìn)而被選擇性地破壞。但含有部分D型氨基酸的多肽則顯示了很強的抵抗蛋白酶降解能力。
DOI | 名稱 | |
---|---|---|
10.1111/j.1471-4159.2007.05052.x | Expression, binding, and signaling properties of CRF2(a) receptors endogenously expressed in human retinoblastoma Y79 cells: passage-dependent regulation of functional receptors | 下載 |
10.1016/j.bbr.2011.03.026 | Wake-promoting effects of orexin: Its independent actions against the background of an impaired corticotropine-releasing hormone receptor system | 下載 |
10.1073/pnas.95.26.15264 | Structural requirements for peptidic antagonists of the corticotropin-releasing factor receptor (CRFR): development of CRFR2beta-selective antisauvagine-30 | 下載 |
多肽H2N-DPhe-His-Leu-Leu-Arg-Lys-Met-Ile-Glu-Ile-Glu-Lys-Gln-Glu-Lys-Glu-Lys-Gln-Gln-Ala-Ala-Asn-Asn-Arg-Leu-Leu-Leu-Asp-Thr-Ile-NH2的合成步驟:
1、合成MBHA樹脂:取若干克的MBHA樹脂(如初始取代度為0.5mmol/g)和1倍樹脂摩爾量的Fmoc-Linker-OH加入到反應(yīng)器中,加入DMF,攪拌使氨基酸完全溶解。再加入樹脂2倍量的DIEPA,攪拌混合均勻。再加入樹脂0.95倍量的HBTU,攪拌混合均勻。反應(yīng)3-4小時后,用DMF洗滌3次。用2倍樹脂體積的10%乙酸酐/DMF 進(jìn)行封端30分鐘。然后再用DMF洗滌3次,甲醇洗滌2次,DCM洗滌2次,再用甲醇洗滌2次。真空干燥12小時以上,得到干燥的樹脂{Fmoc-Linker-MHBA Resin},測定取代度。這里測得取代度為 0.3mmol/g。結(jié)構(gòu)如下圖:
2、脫Fmoc:取1.72g的上述樹脂,用DCM或DMF溶脹20分鐘。用DMF洗滌2遍。加3倍樹脂體積的20%Pip/DMF溶液,鼓氮氣30分鐘,然后2倍樹脂體積的DMF 洗滌5次。得到 H2N-Linker-MBHA Resin 。(此步驟脫除Fmoc基團(tuán),茚三酮檢測為藍(lán)色,Pip為哌啶)。結(jié)構(gòu)圖如下:
3、縮合:取1.55mmol Fmoc-Ile-OH 氨基酸,加入到上述樹脂里,加適當(dāng)DMF溶解氨基酸,再依次加入3.1mmol DIPEA,1.47mmol HBTU。反應(yīng)30分鐘后,取小樣洗滌,茚三酮檢測為無色。用2倍樹脂體積的DMF 洗滌3次樹脂。(洗滌樹脂,去掉殘留溶劑,為下一步反應(yīng)做準(zhǔn)備)。得到Fmoc-Ile-Linker-MBHA Resin。氨基酸:DIPEA:HBTU:樹脂=3:6:2.85:1(摩爾比)。結(jié)構(gòu)圖如下:
4、依次循環(huán)步驟二、步驟三,依次得到
H2N-Ile-Linker-MBHA Resin
Fmoc-Thr(tBu)-Ile-Linker-MBHA Resin
H2N-Thr(tBu)-Ile-Linker-MBHA Resin
Fmoc-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
H2N-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
Fmoc-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
H2N-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
Fmoc-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
H2N-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
Fmoc-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
H2N-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
Fmoc-Arg(Pbf)-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
H2N-Arg(Pbf)-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
Fmoc-Asn(Trt)-Arg(Pbf)-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
H2N-Asn(Trt)-Arg(Pbf)-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
Fmoc-Asn(Trt)-Asn(Trt)-Arg(Pbf)-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
H2N-Asn(Trt)-Asn(Trt)-Arg(Pbf)-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
Fmoc-Ala-Asn(Trt)-Asn(Trt)-Arg(Pbf)-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
H2N-Ala-Asn(Trt)-Asn(Trt)-Arg(Pbf)-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
Fmoc-Ala-Ala-Asn(Trt)-Asn(Trt)-Arg(Pbf)-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
H2N-Ala-Ala-Asn(Trt)-Asn(Trt)-Arg(Pbf)-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
Fmoc-Gln(Trt)-Ala-Ala-Asn(Trt)-Asn(Trt)-Arg(Pbf)-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
H2N-Gln(Trt)-Ala-Ala-Asn(Trt)-Asn(Trt)-Arg(Pbf)-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
Fmoc-Gln(Trt)-Gln(Trt)-Ala-Ala-Asn(Trt)-Asn(Trt)-Arg(Pbf)-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
H2N-Gln(Trt)-Gln(Trt)-Ala-Ala-Asn(Trt)-Asn(Trt)-Arg(Pbf)-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
Fmoc-Lys(Boc)-Gln(Trt)-Gln(Trt)-Ala-Ala-Asn(Trt)-Asn(Trt)-Arg(Pbf)-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
H2N-Lys(Boc)-Gln(Trt)-Gln(Trt)-Ala-Ala-Asn(Trt)-Asn(Trt)-Arg(Pbf)-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
Fmoc-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Gln(Trt)-Ala-Ala-Asn(Trt)-Asn(Trt)-Arg(Pbf)-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
H2N-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Gln(Trt)-Ala-Ala-Asn(Trt)-Asn(Trt)-Arg(Pbf)-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
Fmoc-Lys(Boc)-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Gln(Trt)-Ala-Ala-Asn(Trt)-Asn(Trt)-Arg(Pbf)-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
H2N-Lys(Boc)-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Gln(Trt)-Ala-Ala-Asn(Trt)-Asn(Trt)-Arg(Pbf)-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
Fmoc-Glu(OtBu)-Lys(Boc)-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Gln(Trt)-Ala-Ala-Asn(Trt)-Asn(Trt)-Arg(Pbf)-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
H2N-Glu(OtBu)-Lys(Boc)-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Gln(Trt)-Ala-Ala-Asn(Trt)-Asn(Trt)-Arg(Pbf)-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
Fmoc-Gln(Trt)-Glu(OtBu)-Lys(Boc)-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Gln(Trt)-Ala-Ala-Asn(Trt)-Asn(Trt)-Arg(Pbf)-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
H2N-Gln(Trt)-Glu(OtBu)-Lys(Boc)-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Gln(Trt)-Ala-Ala-Asn(Trt)-Asn(Trt)-Arg(Pbf)-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
Fmoc-Lys(Boc)-Gln(Trt)-Glu(OtBu)-Lys(Boc)-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Gln(Trt)-Ala-Ala-Asn(Trt)-Asn(Trt)-Arg(Pbf)-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
H2N-Lys(Boc)-Gln(Trt)-Glu(OtBu)-Lys(Boc)-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Gln(Trt)-Ala-Ala-Asn(Trt)-Asn(Trt)-Arg(Pbf)-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
Fmoc-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Glu(OtBu)-Lys(Boc)-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Gln(Trt)-Ala-Ala-Asn(Trt)-Asn(Trt)-Arg(Pbf)-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
H2N-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Glu(OtBu)-Lys(Boc)-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Gln(Trt)-Ala-Ala-Asn(Trt)-Asn(Trt)-Arg(Pbf)-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
Fmoc-Ile-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Glu(OtBu)-Lys(Boc)-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Gln(Trt)-Ala-Ala-Asn(Trt)-Asn(Trt)-Arg(Pbf)-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
H2N-Ile-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Glu(OtBu)-Lys(Boc)-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Gln(Trt)-Ala-Ala-Asn(Trt)-Asn(Trt)-Arg(Pbf)-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
Fmoc-Glu(OtBu)-Ile-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Glu(OtBu)-Lys(Boc)-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Gln(Trt)-Ala-Ala-Asn(Trt)-Asn(Trt)-Arg(Pbf)-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
H2N-Glu(OtBu)-Ile-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Glu(OtBu)-Lys(Boc)-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Gln(Trt)-Ala-Ala-Asn(Trt)-Asn(Trt)-Arg(Pbf)-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
Fmoc-Ile-Glu(OtBu)-Ile-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Glu(OtBu)-Lys(Boc)-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Gln(Trt)-Ala-Ala-Asn(Trt)-Asn(Trt)-Arg(Pbf)-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
H2N-Ile-Glu(OtBu)-Ile-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Glu(OtBu)-Lys(Boc)-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Gln(Trt)-Ala-Ala-Asn(Trt)-Asn(Trt)-Arg(Pbf)-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
Fmoc-Met-Ile-Glu(OtBu)-Ile-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Glu(OtBu)-Lys(Boc)-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Gln(Trt)-Ala-Ala-Asn(Trt)-Asn(Trt)-Arg(Pbf)-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
H2N-Met-Ile-Glu(OtBu)-Ile-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Glu(OtBu)-Lys(Boc)-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Gln(Trt)-Ala-Ala-Asn(Trt)-Asn(Trt)-Arg(Pbf)-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
Fmoc-Lys(Boc)-Met-Ile-Glu(OtBu)-Ile-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Glu(OtBu)-Lys(Boc)-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Gln(Trt)-Ala-Ala-Asn(Trt)-Asn(Trt)-Arg(Pbf)-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
H2N-Lys(Boc)-Met-Ile-Glu(OtBu)-Ile-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Glu(OtBu)-Lys(Boc)-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Gln(Trt)-Ala-Ala-Asn(Trt)-Asn(Trt)-Arg(Pbf)-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
Fmoc-Arg(Pbf)-Lys(Boc)-Met-Ile-Glu(OtBu)-Ile-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Glu(OtBu)-Lys(Boc)-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Gln(Trt)-Ala-Ala-Asn(Trt)-Asn(Trt)-Arg(Pbf)-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
H2N-Arg(Pbf)-Lys(Boc)-Met-Ile-Glu(OtBu)-Ile-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Glu(OtBu)-Lys(Boc)-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Gln(Trt)-Ala-Ala-Asn(Trt)-Asn(Trt)-Arg(Pbf)-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
Fmoc-Leu-Arg(Pbf)-Lys(Boc)-Met-Ile-Glu(OtBu)-Ile-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Glu(OtBu)-Lys(Boc)-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Gln(Trt)-Ala-Ala-Asn(Trt)-Asn(Trt)-Arg(Pbf)-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
H2N-Leu-Arg(Pbf)-Lys(Boc)-Met-Ile-Glu(OtBu)-Ile-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Glu(OtBu)-Lys(Boc)-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Gln(Trt)-Ala-Ala-Asn(Trt)-Asn(Trt)-Arg(Pbf)-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
Fmoc-Leu-Leu-Arg(Pbf)-Lys(Boc)-Met-Ile-Glu(OtBu)-Ile-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Glu(OtBu)-Lys(Boc)-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Gln(Trt)-Ala-Ala-Asn(Trt)-Asn(Trt)-Arg(Pbf)-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
H2N-Leu-Leu-Arg(Pbf)-Lys(Boc)-Met-Ile-Glu(OtBu)-Ile-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Glu(OtBu)-Lys(Boc)-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Gln(Trt)-Ala-Ala-Asn(Trt)-Asn(Trt)-Arg(Pbf)-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
Fmoc-His(Trt)-Leu-Leu-Arg(Pbf)-Lys(Boc)-Met-Ile-Glu(OtBu)-Ile-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Glu(OtBu)-Lys(Boc)-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Gln(Trt)-Ala-Ala-Asn(Trt)-Asn(Trt)-Arg(Pbf)-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
H2N-His(Trt)-Leu-Leu-Arg(Pbf)-Lys(Boc)-Met-Ile-Glu(OtBu)-Ile-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Glu(OtBu)-Lys(Boc)-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Gln(Trt)-Ala-Ala-Asn(Trt)-Asn(Trt)-Arg(Pbf)-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
Fmoc-DPhe-His(Trt)-Leu-Leu-Arg(Pbf)-Lys(Boc)-Met-Ile-Glu(OtBu)-Ile-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Glu(OtBu)-Lys(Boc)-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Gln(Trt)-Ala-Ala-Asn(Trt)-Asn(Trt)-Arg(Pbf)-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin
以上中間結(jié)構(gòu),均可在專肽生物多肽計算器-多肽結(jié)構(gòu)計算器中,一鍵畫出。
最后再經(jīng)過步驟二得到 H2N-DPhe-His(Trt)-Leu-Leu-Arg(Pbf)-Lys(Boc)-Met-Ile-Glu(OtBu)-Ile-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Glu(OtBu)-Lys(Boc)-Glu(OtBu)-Lys(Boc)-Gln(Trt)-Gln(Trt)-Ala-Ala-Asn(Trt)-Asn(Trt)-Arg(Pbf)-Leu-Leu-Leu-Asp(OtBu)-Thr(tBu)-Ile-Linker-MBHA Resin,結(jié)構(gòu)如下:
5、切割:6倍樹脂體積的切割液(或每1g樹脂加8ml左右的切割液),搖床搖晃 2小時,過濾掉樹脂,用冰無水乙醚沉淀濾液,并用冰無水乙醚洗滌沉淀物3次,最后將沉淀物放真空干燥釜中,常溫干燥24小試,得到粗品H2N-DPhe-His-Leu-Leu-Arg-Lys-Met-Ile-Glu-Ile-Glu-Lys-Gln-Glu-Lys-Glu-Lys-Gln-Gln-Ala-Ala-Asn-Asn-Arg-Leu-Leu-Leu-Asp-Thr-Ile-NH2。結(jié)構(gòu)圖見產(chǎn)品結(jié)構(gòu)圖。
切割液選擇: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%
(前兩種適合沒有容易氧化的氨基酸,例如Trp、Cys、Met。第三種適合幾乎所有的序列。)
6、純化凍干:使用液相色譜純化,收集目標(biāo)峰液體,進(jìn)行凍干,獲得蓬松的粉末狀固體多肽。不過這時要取小樣復(fù)測下純度 是否目標(biāo)純度。
7、最后總結(jié):
杭州專肽生物技術(shù)有限公司(ALLPEPTIDE http://amynixphotography.com)主營定制多肽合成業(yè)務(wù),提供各類長肽,短肽,環(huán)肽,提供各類修飾肽,如:熒光標(biāo)記修飾(CY3、CY5、CY5.5、CY7、FAM、FITC、Rhodamine B、TAMRA等),功能基團(tuán)修飾肽(疊氮、炔基、DBCO、DOTA、NOTA等),同位素標(biāo)記肽(N15、C13),訂書肽(Stapled Peptide),脂肪酸修飾肽(Pal、Myr、Ste),磷酸化修飾肽(P-Ser、P-Thr、P-Tyr),環(huán)肽(酰胺鍵環(huán)肽、一對或者多對二硫鍵環(huán)),生物素標(biāo)記肽,PEG修飾肽,甲基化修飾肽等。
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