400-998-5282
專注多肽 服務(wù)科研
編號: | 181536 |
中文名稱: | 腸抑素Enterostatin, porcine, rat |
CAS號: | 117137-85-6 |
單字母: | H2N-VPDPR-OH |
三字母: | H2N N端氨基 -Val纈氨酸 -Pro脯氨酸 -Asp天冬氨酸 -Pro脯氨酸 -Arg精氨酸 -OHC端羧基 |
氨基酸個(gè)數(shù): | 5 |
分子式: | C25H42N8O8 |
平均分子量: | 582.65 |
精確分子量: | 582.31 |
等電點(diǎn)(PI): | 10.56 |
pH=7.0時(shí)的凈電荷數(shù): | 0.98 |
堿性基團(tuán)個(gè)數(shù): | 非常親水 |
平均親水性: | 1.5 |
疏水性值: | -1.4 |
外觀與性狀: | 白色粉末狀固體 |
閃點(diǎn): | 0 M-1cm-1 |
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或其它 |
儲存條件: | 負(fù)80℃至負(fù)20℃ |
標(biāo)簽: | 肥胖研究 腸抑素(Enterostatins) |
Definition
Enterostatin is a pentapeptide generated by the action of trypsin on procolipase in the intestinal lumen. Pharmacologic studies have suggested a role for these peptides in appetite regulation and insulin secretion 1.
Related Peptides
Enterostatins belong to a family of peptides (e.g., Val-Pro-Asp-Pro-Arg, VPDPR; Ala-Pro-Gly-Pro-Arg, APGPR; and Val-Pro-Gly-Pro-Arg, VPGPR) derived from the tryptic cleavage of amino-terminal pentapeptide from procolipase 1.
Discovery
In 1993, Mei et al., identified the peptapeptide obtained from the tryptic cleavage of the N-terminal of procolipase and named it enterostatin 2.
Structural Characteristics
Enterostatin structure is highly conserved in evolution, with an amino acid sequence of XPXPR. Three enterostatin sequences, Val-Pro-Asp-Pro-Arg (VPDPR), Val-Pro-Gly-Pro-Arg (VPGPR), and Ala-Pro-Gly-Pro-Arg (APGPR), have been studied extensively and shown to be almost equally effective in their ability to decrease dietary fat preference 3. Modifications in the N-terminus of VPDPR abrogate biological function, other modifications result in retention of biological activity. The importance of the N-terminal residue of enterostatin was investigated by replacing the L-valine with (i) D-valine, (ii) L-tyrosine or (iii) addition of an L-tyrosine to generate the hexapeptide, YVPDPR. Replacement of the N-terminal L-valine residue with D-valine abrogated the insulin inhibitory activity of the peptide. This demonstrates that the N-terminal valine may be necessary for biological function and is consistent with a previous report showing a lack of effect of D-VPDPR on high-fat food consumption 4.
Mode of Action
A high-fat diet easily promotes hyperphagia giving an impression of an uncontrolled process. Fat digestion itself however provides control of fat intake through the digestion, carried out by pancreatic lipase and its protein cofactor colipase, and through enterostatin, released from procolipase during fat digestion 5. Neural mechanisms arising from the intestine involving vagal afferent nerves may be important for the early satiety mechanism induced by enterostatin during a meal 6. Enterostatin acts through an opioid pathway, rather as a µ-antagonist acting antagonistic to ß-casomorphin. In its mechanism of action enterostatin gives an exothermic reaction, while the opiate has an endothermic reaction. The decreased thermogenesis by opiates could explain an increased appetite and body weight, while an increased thermogenesis by enterostatin could explain a reduction in appetite and loss of body weight. The hypocholesterolemic effects of APGPR and VPDPR are mediated by a CCK1 receptor-dependent mechanism 7.
Functions
Enterostatin released from the exocrine pancreas and gastrointestinal tract, selectively inhibits fat intake through activation of an afferent vagal signaling pathway 8. Recent studies indicate that enterostatin modulates insulin release in response to a variety of agents that affect first phase as well as the second phase of insulin release 4.
References
1. Imamura M, Sumar N, Hermon-Taylor J, Robertson HJ, Prasad C (1998). Distribution and characterization of enterostatin-like immunoreactivity in human cerebrospinal fluid. Peptides, 19(8):1385-1391.
2. Mei J, Bowyer RC, Jehanli AMT, Patel G, Erlanson-Albertsson C (1993). Identification of enterostatin, the pancreatic procolipase activation peptide in the intestine of rat. Pancreas, 8:488-493.
3. Prasad C, Imamura M, Debata C, Svec F, Sumar N, Hermon-Taylor J (1999). Hyper- enterostatinemia in Premenopausal Obese Women. J Clin Endocrinol Metab., 84(3):937-941.
4. Tadayyon M, Liou S, Briscoe CP, Badman G, Eggleston DS, Arch JRS, York DA (2002). Structure-function studies on enterostatin inhibition of insulin release. Int J Diabetes & Metabolism., 10:14-21.
5. Berger K, Winzell MS, Mei J, Erlanson-Albertsson C (2004). Enterostatin and its target mechanisms during regulation of fat intake. Physiol Behav., 83(4):623-630.
6. Mei J, Sörhede-Winzell M, Erlanson-Albertsson C (2002). Plasma Enterostatin: Identification and Release in Rats in Response to a Meal. Obes Res., 10(7):688–694.
7. Takenaka Y, Shimano T, Mori T, Hou IC, Ohinata K, Yoshikawa M. (2008). Enterostatin reduces serum cholesterol levels by way of a CCK1 receptor-dependent mechanism. Peptides.,29(12):2175-2178.
8. Lin L, Thomas SR, Kilroy G, Schwartz GJ, York DA (2003). Enterostatin inhibition of dietary fat intake is dependent on CCK-A receptors Am J Physiol Regul Integr Comp Physiol., 285(2):321-328.
多肽H2N-Val-Pro-Asp-Pro-Arg-COOH的合成步驟:
1、合成CTC樹脂:稱取2.78g CTC Resin(如初始取代度約為0.6mmol/g)和2.0mmol Fmoc-Arg(Pbf)-OH于反應(yīng)器中,加入適量DCM溶解氨基酸(需要注意,此時(shí)CTC樹脂體積會增大好幾倍,避免DCM溶液過少),再加入5.0mmol DIPEA(Mw:129.1,d:0.740g/ml),反應(yīng)2-3小時(shí)后,可不抽濾溶液,直接加入1ml的HPLC級甲醇,封端半小時(shí)。依次用DMF洗滌2次,甲醇洗滌1次,DCM洗滌一次,甲醇洗滌一次,DCM洗滌一次,DMF洗滌2次(這里使用甲醇和DCM交替洗滌,是為了更好地去除其他溶質(zhì),有利于后續(xù)反應(yīng))。得到 Fmoc-Arg(Pbf)-CTC Resin。結(jié)構(gòu)圖如下:
2、脫Fmoc:加3倍樹脂體積的20%Pip/DMF溶液,鼓氮?dú)?0分鐘,然后2倍樹脂體積的DMF 洗滌5次。得到 H2N-Arg(Pbf)-CTC 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5.0mmol Fmoc-Pro-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10.01mmol DIPEA,4.75mmol 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-Pro-Arg(Pbf)-CTC 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-Pro-Arg(Pbf)-CTC Resin
Fmoc-Asp(OtBu)-Pro-Arg(Pbf)-CTC Resin
H2N-Asp(OtBu)-Pro-Arg(Pbf)-CTC Resin
Fmoc-Pro-Asp(OtBu)-Pro-Arg(Pbf)-CTC Resin
H2N-Pro-Asp(OtBu)-Pro-Arg(Pbf)-CTC Resin
Fmoc-Val-Pro-Asp(OtBu)-Pro-Arg(Pbf)-CTC Resin
以上中間結(jié)構(gòu),均可在專肽生物多肽計(jì)算器-多肽結(jié)構(gòu)計(jì)算器中,一鍵畫出。
最后再經(jīng)過步驟二得到 H2N-Val-Pro-Asp(OtBu)-Pro-Arg(Pbf)-CTC 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(shí),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-Val-Pro-Asp-Pro-Arg-COOH。結(jié)構(gòu)圖見產(chǎn)品結(jié)構(gòu)圖。
切割液選擇:1)TFA:H2O=95%:5%、TFA:H2O=97.5%:2.5%
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(shí)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://tsjxdd.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
以上所有內(nèi)容,為專肽生物原創(chuàng)內(nèi)容,請勿發(fā)布到其他網(wǎng)站上。