Amylin (20-29) (or hIAPP 20-29) forms fibrils that are ultrastructurally identical to amyloid fibrils seen in pancreatic islets. The region SNNFGAILSS appears to be the most important amyloidogenic sequence of hIAPP (human islet amyloid polypeptide).
编号：192725
CAS号：118068-30-7
单字母：H2N-SNNFGAILSS-OH
| 编号: | 192725 |
| 中文名称: | 胰淀素Amylin(20-29), human |
| 英文名: | Amylin(20-29), human |
| CAS号: | 118068-30-7 |
| 单字母: | H2N-SNNFGAILSS-OH |
| 三字母: | H2N N端氨基 -Ser丝氨酸 -Asn天冬酰胺 -Asn天冬酰胺 -Phe苯丙氨酸 -Gly甘氨酸 -Ala丙氨酸 -Ile异亮氨酸 -Leu亮氨酸 -Ser丝氨酸 -Ser丝氨酸 -OHC端羧基 |
| 氨基酸个数: | 10 |
| 分子式: | C43H68N12O16 |
| 平均分子量: | 1009.07 |
| 精确分子量: | 1008.49 |
| 等电点(PI): | - |
| pH=7.0时的净电荷数: | 0.97 |
| 平均亲水性: | -0.58888888888889 |
| 疏水性值: | 0.31 |
| 外观与性状: | 白色粉末状固体 |
| 消光系数: | - |
| 来源: | 人工化学合成，仅限科学研究使用，不得用于人体。 |
| 纯度: | 95%、98% |
| 盐体系: | 可选TFA、HAc、HCl或其它 |
| 储存条件: | 负80℃至负20℃ |
| 标签: | 胰淀素(Amylin) |
Amylin (20-29) (or hIAPP 20-29) forms fibrils that are ultrastructurally identical to amyloid fibrils seen in pancreatic islets. The region SNNFGAILSS appears to be the most important amyloidogenic sequence of hIAPP (human islet amyloid polypeptide).
Definition
Amylin or Islet amyloid polypeptide (IAPP), a 37-amino acid peptide is secreted by beta-islet cells of the pancreas and a major component of the amyloid deposits in persons with type 2 diabetes mellitus. Amylin may be referred to as insulin’s “fraternal twin” as it is constitutively expressed with insulin in response to elevations of plasma glucose.
Discovery
The knowledge of occurrence of amyloid deposits in islets of Langerhans, major pathologic feature of diabetics has been known for a century. But, the discovery of amylin as a major component of amyloid deposits was by two independent groups in 19871, 2.
Structural characteristics
The human amylin has an amino acid sequence KCNTATCATQRLANFLVHSSNNFGAILSSTNVGSNTY, with a disulfide bridge between cysteine residues 2 and 7. The features, amidated C-terminus and the disulfide bridge are necessary for the full biological activity of amylin3. Amylin amino acid sequence is 46% and 43% identical to those of the calcitonin gene-related neuropeptides CGRP-2 and CGRP-1. The (20-29) fragment of amylin is critical to the pathogenesis of islet amyloid4.
Mechanism of action
Amylin is synthesized, packaged within the golgi apparatus and secreted within the secretory granule by the islet beta cell. They have binding sites within the renal cortex in the area of the juxtaglomerular apparatus and it activates the rennin angiotensin aldosterone system. It also acts upon the circulatory system by inhibiting the secretion of the atrial natriuretic peptide (ANP)5.
Function
Amylin inhibits gastric emptying and is important in controlling and delaying the rate of meal derived glucose. It inhibits hepatic release and production of glucose in the postprandial period. They also have been shown to inhibit glucagon secretion and somatostatin. Amylin causes vasodilatation by dilating the non-striated muscles of the blood vessels. It is also known to increase thirst level which indicates it has an action within the central nervous system6.
References
1. Cooper GJ, Willis AC, Clark A, Turner RC, Sim RB, Reid KB (1987). Purification and characterization of a peptide from amyloid-rich pancreases of type 2 diabetic patients. Proc Natl Acad Sci., 84(23):8628-32.
2. Westermark P, Wernstedt C, O'Brien TD, Hayden DW, Johnson KH (1987). Islet amyloid in type 2 human diabetes mellitus and adult diabetic cats contains a novel putative polypeptide hormone. Am J Pathol, 127(3):414-417.
3. Roberts AN, Leighton B, Todd JA, Cockburn D, Schofield PN, Sutton R, Holt S, Boyd Y, Day AJ, Foot EA, et al,(1989). Molecular and functional characterization of amylin, a peptide associated with type 2 diabetes mellitus. Proc Natl Acad Sci, 86(24):9662-9666.
4. Guidobono F, Pagani F, Ticozzi C, Sibilia V, Pecile A, Netti C (1997). Protection by amylin of gastric erosions induced by indomethacin or ethanol in rats. Br J Pharmacol, 120(4):581-586.
5. Piao FL, Cao C, Han JH, Kim SZ, Cho KW, Kim SH (2004). Amylin-induced suppression of ANP secretion through receptors for CGRP1 and salmon calcitonin. Regul Pept, 117: 59-166.
6. Hayden MR, Tyagi SC (2002). Islet redox stress: the manifold toxicities of insulin resistance, metabolic syndrome and amylin derived islet amyloid in type 2 diabetes mellitus. Journal of the Pancrease, 3(4): 86-108.
| DOI | 名称 | |
|---|---|---|
| 10.1016/S0006-3495(03)70068-X | Micelle formation by a fragment of human islet amyloid polypeptide | 下载 |
| 10.1177/030098589303000401 | Islet amyloid polypeptide: a review of its biology and potential roles in the pathogenesis of diabetes mellitus | 下载 |
多肽H2N-Ser-Asn-Asn-Phe-Gly-Ala-Ile-Leu-Ser-Ser-COOH的合成步骤：
1、合成CTC树脂：称取0.56g CTC Resin（如初始取代度约为0.84mmol/g）和0.56mmol Fmoc-Ser(tBu)-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1.41mmol 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-Ser(tBu)-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-Ser(tBu)-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1.41mmol Fmoc-Ser(tBu)-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2.82mmol DIPEA，1.34mmol 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-Ser(tBu)-Ser(tBu)-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-Ser(tBu)-Ser(tBu)-CTC Resin
Fmoc-Leu-Ser(tBu)-Ser(tBu)-CTC Resin
H2N-Leu-Ser(tBu)-Ser(tBu)-CTC Resin
Fmoc-Ile-Leu-Ser(tBu)-Ser(tBu)-CTC Resin
H2N-Ile-Leu-Ser(tBu)-Ser(tBu)-CTC Resin
Fmoc-Ala-Ile-Leu-Ser(tBu)-Ser(tBu)-CTC Resin
H2N-Ala-Ile-Leu-Ser(tBu)-Ser(tBu)-CTC Resin
Fmoc-Gly-Ala-Ile-Leu-Ser(tBu)-Ser(tBu)-CTC Resin
H2N-Gly-Ala-Ile-Leu-Ser(tBu)-Ser(tBu)-CTC Resin
Fmoc-Phe-Gly-Ala-Ile-Leu-Ser(tBu)-Ser(tBu)-CTC Resin
H2N-Phe-Gly-Ala-Ile-Leu-Ser(tBu)-Ser(tBu)-CTC Resin
Fmoc-Asn(Trt)-Phe-Gly-Ala-Ile-Leu-Ser(tBu)-Ser(tBu)-CTC Resin
H2N-Asn(Trt)-Phe-Gly-Ala-Ile-Leu-Ser(tBu)-Ser(tBu)-CTC Resin
Fmoc-Asn(Trt)-Asn(Trt)-Phe-Gly-Ala-Ile-Leu-Ser(tBu)-Ser(tBu)-CTC Resin
H2N-Asn(Trt)-Asn(Trt)-Phe-Gly-Ala-Ile-Leu-Ser(tBu)-Ser(tBu)-CTC Resin
Fmoc-Ser(tBu)-Asn(Trt)-Asn(Trt)-Phe-Gly-Ala-Ile-Leu-Ser(tBu)-Ser(tBu)-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-Ser(tBu)-Asn(Trt)-Asn(Trt)-Phe-Gly-Ala-Ile-Leu-Ser(tBu)-Ser(tBu)-CTC 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-Ser-Asn-Asn-Phe-Gly-Ala-Ile-Leu-Ser-Ser-COOH。结构图见产品结构图。
切割液选择：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：
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