神经激素异源催产素 GFKNVEMMTARGF-酰胺已从成年烟草角虫 Manduca sexta 的头部提取物中分离出来。它在体外强烈刺激保幼激素从语料库中释放。
编号：186128
CAS号：120928-88-3
单字母：H2N-GFKNVEMMTARGF-NH2
| 编号: | 186128 |
| 中文名称: | 抑咽侧体神经肽Allatotropin |
| 英文名: | Allatotropin |
| 英文同义词: | Allatotropin, Mas - AT |
| CAS号: | 120928-88-3 |
| 单字母: | H2N-GFKNVEMMTARGF-NH2 |
| 三字母: | H2N N端氨基 -Gly甘氨酸 -Phe苯丙氨酸 -Lys赖氨酸 -Asn天冬酰胺 -Val缬氨酸 -Glu谷氨酸 -Met甲硫氨酸 -Met甲硫氨酸 -Thr苏氨酸 -Ala丙氨酸 -Arg精氨酸 -Gly甘氨酸 -Phe苯丙氨酸 -NH2C端酰胺化 |
| 氨基酸个数: | 13 |
| 分子式: | C65H103N19O17S2 |
| 平均分子量: | 1486.76 |
| 精确分子量: | 1485.72 |
| 等电点(PI): | 11.66 |
| pH=7.0时的净电荷数: | 1.98 |
| 平均亲水性: | -0.072727272727273 |
| 疏水性值: | -0.12 |
| 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: | 抑咽侧体神经肽(Allatostatin) |
The neurohormone allatotropin, GFKNVEMMTARGF-amide, has been isolated from head extracts of the adult tobacco hornworm, Manduca sexta. It strongly stimulates the release of juvenile hormone from the corpora allata in vitro.
Definition
Allatostatins (ASTs) are pleiotropic neuropeptide hormones in insects and crustacean. There major function in the insect is the inhibition of juvenile hormone synthesis by the corpora allata and reduce their food intake1.
Discovery
The first identified ASTs were isolated from brain-retro cerebral complexes of the cockroach Diploptera punctata2.
Classification
ASTs encompass a group of three families determined by consensus sequences, the cockroach type representing the FGLa family, the cricket type representing the W(X)6 Wa family and the PISCF family 1.
Structural Characteristics
AST’s are 8-13 amino acids long, are amidated, and show sequence similarity, including a 3-amino acid sequence at the C-terminal end that is common to all four peptides. The peptide sequences are as follows: allatostatin-1, Ala-Pro-Ser-Gly-Ala-Gln-Arg-Leu-Tyr-Gly-Phe-Gly-Leu-NH2; allatostatin-2, Gly-Asp-Gly-Arg-Leu-Tyr-Ala-Phe-Gly-Leu-NH2; allatostatin-3, Gly-Gly-Ser-Leu-Tyr-Ser-Phe-Gly-Leu-NH2; and allatostatin-4, Asp-Arg-Leu-Tyr-Ser-Phe-Gly-Leu-NH23.
Mode of action
In mammalian cells, allatostatin bind to AST receptors to open G-protein coupled inward rectifying potassium channels, resulting in reduced membrane potential and input resistance4.
References
1. Stay B and Tobe SS (2007). "The role of allatostatins in juvenile hormone synthesis in insects and crustaceans". Annu. Rev. Entomo., 52: 277–99.
2. Pratt GE, Farnsworth DE, Fok KF, Siegel NR, McCormack AL, Shabanowitz J, Hunt DF, Feyereisen R (1991). Identity of a second type of allatostatin from cockroach brains: an octadecapeptide amide with a tyrosine-rich address sequence. Proc Natl Acad Sci., 88(6): 2412–2416.
3. Woodhead AP, Stay B, Seidel SL, Khan MA, Tobe SS (1989). Primary structure of four allatostatins: neuropeptide inhibitors of juvenile hormone synthesis. Proc Natl Acad Sci., 86(15):5997-6001.
4. Tan EM, Yamaguchi Y, Horwitz GD, Gosgnach S, Lein ES, Goulding M, Albright TD, Callaway EM (2006). Selective and quickly reversible inactivation of mammalian neurons in vivo uses the Drosophila allatostatin receptor. Neuron, 1:57–170.
| DOI | 名称 | |
|---|---|---|
| 10.1016/s0196-9781(01)00660-x | Effects of allatotropin and allatostatin on in vitro production of juvenile hormones by the corpora allata of virgin females of the moths of Heliothis virescens and Manduca sexta | 下载 |
| 10.1016/s0965-1748(03)00005-5 | Molecular characterisation of cDNAs from the fall armyworm Spodoptera frugiperda encoding Manduca sexta allatotropin and allatostatin preprohormone peptides | 下载 |
| 10.1016/s0022-1910(03)00017-9 | Myotropic effect of helicokinins, tachykinin-related peptides and Manduca sexta allatotropin on the gut of Heliothis virescens (Lepidoptera: Noctuidae) | 下载 |
| 10.1016/j.regpep.2004.05.015 | Characterization of a novel peptide with allatotropic activity in the fall armyworm Spodoptera frugiperda | 下载 |
| 10.1002/dneu.20381 | Direct peptide profiling of lateral cell groups of the antennal lobes of Manduca sexta reveals specific composition and changes in neuropeptide expression during development | 下载 |
| 10.1126/science.243.4897.1481 | Identification of an allatotropin from adult manduca sexta | 下载 |
| 10.1002/arch.20332 | Control of the release of digestive enzymes in the larvae of the fall armyworm, Spodoptera frugiperda | 下载 |
| 10.1016/j.ibmb.2011.06.002 | Isolation and functional characterization of an allatotropin receptor from Manduca sexta | 下载 |
多肽H2N-Gly-Phe-Lys-Asn-Val-Glu-Met-Met-Thr-Ala-Arg-Gly-Phe-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2.23g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2.01mmol Fmoc-Phe-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4.01mmol DIPEA，1.91mmol 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-Phe-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-Phe-Linker-MBHA Resin
Fmoc-Gly-Phe-Linker-MBHA Resin
H2N-Gly-Phe-Linker-MBHA Resin
Fmoc-Arg(Pbf)-Gly-Phe-Linker-MBHA Resin
H2N-Arg(Pbf)-Gly-Phe-Linker-MBHA Resin
Fmoc-Ala-Arg(Pbf)-Gly-Phe-Linker-MBHA Resin
H2N-Ala-Arg(Pbf)-Gly-Phe-Linker-MBHA Resin
Fmoc-Thr(tBu)-Ala-Arg(Pbf)-Gly-Phe-Linker-MBHA Resin
H2N-Thr(tBu)-Ala-Arg(Pbf)-Gly-Phe-Linker-MBHA Resin
Fmoc-Met-Thr(tBu)-Ala-Arg(Pbf)-Gly-Phe-Linker-MBHA Resin
H2N-Met-Thr(tBu)-Ala-Arg(Pbf)-Gly-Phe-Linker-MBHA Resin
Fmoc-Met-Met-Thr(tBu)-Ala-Arg(Pbf)-Gly-Phe-Linker-MBHA Resin
H2N-Met-Met-Thr(tBu)-Ala-Arg(Pbf)-Gly-Phe-Linker-MBHA Resin
Fmoc-Glu(OtBu)-Met-Met-Thr(tBu)-Ala-Arg(Pbf)-Gly-Phe-Linker-MBHA Resin
H2N-Glu(OtBu)-Met-Met-Thr(tBu)-Ala-Arg(Pbf)-Gly-Phe-Linker-MBHA Resin
Fmoc-Val-Glu(OtBu)-Met-Met-Thr(tBu)-Ala-Arg(Pbf)-Gly-Phe-Linker-MBHA Resin
H2N-Val-Glu(OtBu)-Met-Met-Thr(tBu)-Ala-Arg(Pbf)-Gly-Phe-Linker-MBHA Resin
Fmoc-Asn(Trt)-Val-Glu(OtBu)-Met-Met-Thr(tBu)-Ala-Arg(Pbf)-Gly-Phe-Linker-MBHA Resin
H2N-Asn(Trt)-Val-Glu(OtBu)-Met-Met-Thr(tBu)-Ala-Arg(Pbf)-Gly-Phe-Linker-MBHA Resin
Fmoc-Lys(Boc)-Asn(Trt)-Val-Glu(OtBu)-Met-Met-Thr(tBu)-Ala-Arg(Pbf)-Gly-Phe-Linker-MBHA Resin
H2N-Lys(Boc)-Asn(Trt)-Val-Glu(OtBu)-Met-Met-Thr(tBu)-Ala-Arg(Pbf)-Gly-Phe-Linker-MBHA Resin
Fmoc-Phe-Lys(Boc)-Asn(Trt)-Val-Glu(OtBu)-Met-Met-Thr(tBu)-Ala-Arg(Pbf)-Gly-Phe-Linker-MBHA Resin
H2N-Phe-Lys(Boc)-Asn(Trt)-Val-Glu(OtBu)-Met-Met-Thr(tBu)-Ala-Arg(Pbf)-Gly-Phe-Linker-MBHA Resin
Fmoc-Gly-Phe-Lys(Boc)-Asn(Trt)-Val-Glu(OtBu)-Met-Met-Thr(tBu)-Ala-Arg(Pbf)-Gly-Phe-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-Gly-Phe-Lys(Boc)-Asn(Trt)-Val-Glu(OtBu)-Met-Met-Thr(tBu)-Ala-Arg(Pbf)-Gly-Phe-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-Gly-Phe-Lys-Asn-Val-Glu-Met-Met-Thr-Ala-Arg-Gly-Phe-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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