Adrenomedullin (16-31), human TFA 是人腎上腺髓質(zhì)素 (hADM) 的 16-31 氨基酸片段。腎上腺髓質(zhì)素對(duì) CGRP1 受體具有明顯的親和力。Adrenomedullin (16-31), human TFA 對(duì)大鼠全身血管具有升壓活性，但對(duì)貓沒(méi)有作用活性。
Adrenomedullin (16-31), human TFA is amino acid residues 16-31 fragment of human adrenomedullin (hADM). Adrenomedullin has appreciable affinity for the CGRP1 receptor. Adrenomedullin (16-31), human TFA possesses pressor activity in the systemic vascular bed of the rat, but not the cat[1].

二硫鍵廣泛存在與蛋白結(jié)構(gòu)中，對(duì)穩(wěn)定蛋白結(jié)構(gòu)具有非常重要的意義，二硫鍵一般是通過(guò)序列中的2個(gè)Cys的巰基，經(jīng)氧化形成。
 

形成二硫鍵的方法很多：空氣氧化法，DMSO氧化法，過(guò)氧化氫氧化法等。
 

二硫鍵的合成過(guò)程，  可以通過(guò)Ellman檢測(cè)以及HPLC檢測(cè)方法對(duì)其反應(yīng)進(jìn)程進(jìn)行監(jiān)測(cè)。  
   

如果多肽中只含有1對(duì)Cys，那二硫鍵的形成是簡(jiǎn)單的。多肽經(jīng)固相或液相合成，然后在pH8-9的溶液中進(jìn)行氧化。      
 

當(dāng)需要形成2對(duì)或2對(duì)以上的二硫鍵時(shí)，合成過(guò)程則相對(duì)復(fù)雜。盡管二硫鍵的形成通常是在合成方案的最后階段完成，但有時(shí)引入預(yù)先形成的二硫化物是有利于連合或延長(zhǎng)肽鏈的。通常采用的巰基保護(hù)基有trt, Acm, Mmt, tBu, Bzl, Mob, Tmob等多種基團(tuán)。我們分別列出兩種以2-Cl樹(shù)脂和Rink樹(shù)脂為載體合成的多肽上多對(duì)二硫鍵形成路線：
 

二硫鍵反應(yīng)條件選擇    
 

 二硫鍵即為蛋白質(zhì)或多肽分子中兩個(gè)不同位點(diǎn)Cys的巰基（-SH）被氧化形成的S-S共價(jià)鍵。 一條肽鏈上不同位置的氨基酸之間形成的二硫鍵，可以將肽鏈折疊成特定的空間結(jié)構(gòu)。多肽分 子通常分子量較大，空間結(jié)構(gòu)復(fù)雜，結(jié)構(gòu)中形成二硫鍵時(shí)要求兩個(gè)半胱氨酸在空間距離上接近。 此外，多肽結(jié)構(gòu)中還原態(tài)的巰基化學(xué)性質(zhì)活潑，容易發(fā)生其他的副反應(yīng)，而且肽鏈上其他側(cè)鏈 也可能會(huì)發(fā)生一系列修飾，因此，肽鏈進(jìn)行修飾所選取的氧化劑和氧化條件是反應(yīng)的關(guān)鍵因素， 反應(yīng)機(jī)理也比較復(fù)雜，既可能是自由基反應(yīng)，也可能是離子反應(yīng)。      

反應(yīng)條件有多種選擇，比如空氣氧化，DMSO氧化等溫和的氧化過(guò)程，也可以采用H2O2，I2， 汞鹽等激烈的反應(yīng)條件。
 

空氣氧化法： 空氣氧化法形成二硫鍵是多肽合成中最經(jīng)典的方法，通常是將巰基處于還原態(tài)的多肽溶于水中，在近中性或弱堿性條件下（PH值6.5-10），反應(yīng)24小時(shí)以上。為了降低分子之間二硫鍵形成的可能，該方法通常需要在低濃度條件下進(jìn)行。
 

碘氧化法：將多肽溶于25%的甲醇水溶液或30%的醋酸水溶液中，逐滴滴加10-15mol/L的碘進(jìn)行氧化，反應(yīng)15-40min。當(dāng)肽鏈中含有對(duì)碘比較敏感的Tyr、Trp、Met和His的殘基時(shí)，氧化條件要控制的更精確，氧化完后，立即加入維生素C或硫代硫酸鈉除去過(guò)量的碘。 當(dāng)序列中有兩對(duì)或多對(duì)二硫鍵需要成環(huán)時(shí)，通常有兩種情況：
 

自然隨機(jī)成環(huán):       序列中的Cys之間隨機(jī)成環(huán)，與一對(duì)二硫鍵成環(huán)條件相似；
 

定點(diǎn)成環(huán)：       定點(diǎn)成環(huán)即序列中的Cys按照設(shè)計(jì)要求形成二硫鍵，反應(yīng)過(guò)程相對(duì)復(fù)雜。在 固相合成多肽之前，需要提前設(shè)計(jì)幾對(duì)二硫鍵形成的順序和方法路線，選擇不同的側(cè)鏈 巰基保護(hù)基，利用其性質(zhì)差異，分步氧化形成兩對(duì)或多對(duì)二硫鍵。       通常采用的巰基保護(hù) 基有trt, Acm, Mmt, tBu, Bzl, Mob, Tmob等多種基團(tuán)。

Definition
Adrenomedullin (AM) is a pluripotent peptide and a hypotensive substance extracted from human adrenal tumour. Due to its origin of discovery, i.e. the medulla of the adrenal gland, the peptide is named adrenomedullin.
Discovery
AM was initially isolated from phaechromcytoma cells in 1993 by Kitmura K and his associates1.

Classification
AM is a member of the calcitonin family of peptides. In teleost fish, AM forms an independent subfamily consisting of five members viz. (AM1–AM5). This teleost AM family comprises three groups, AM1/AM4, AM2/AM3, and AM5 2,3.

Structural Characteristics
The peptide consists of 52 amino acids with a 6-member ring structure linked by a disulfide bond between amino acid 16 and 21 and amidated-COOH terminal4. It has 27 % homology with the calcitonin gene-related peptide (CGRP).

Mechanism of action
AM peptides act through specific receptors in the plasma membrane to activate adenylate cyclase activity and modulate Ca2+ flux in the target cells. The intracellular free Ca2+ increases on the activation of phospholipase C and formation of inositol 1, 4, 5-trisphosphate in the endothelial cells. The intracellular increase of Ca2+ activates endothelial nitric oxide synthase which leads to vascular relaxation5.

Function
AM is the most potent endogenous vasodilatory peptide found in the body6. They increase the tolerance of cells to oxidative stress, hypoxic injury and angiogenesis. It plays an important role in neurotransmission and ovarian function and in kidney, it acts as a diuretic and natriuretic7. AM is considered to play an important endocrine role in various tissues in maintaining electrolyte and fluid homeostasis8. It is used in the diagnosis and treatment of preeclampsia, type II diabetic patients and to promote fetal growth. They also play an important role in the regulation of insulin secretion and blood glucose metabolism.

References

1.     Kitamura K, Kangawa K, Kawamoto M, Ichiki Y, Nakamura S, Matsuo H, Eto T (1993). Adrenomedullin: a novel hypotensive peptide isolated from human pheochromocytoma. Biochem Biophys Res Commun., 192 (2):553-560.

2.     Ogoshi M, Nobata S, and Takei Y (2008). Potent osmoregulatory actions of homologous adrenomedullins administered peripherally and centrally in eels. Am J Physiol Regul Integr Comp Physiol, 295: 2075-2083.

3.     Ogoshi M, Inoue K, Naruse K, Takei Y (2006). Evolutionary history of the calcitonin gene-related peptide family in vertebrates revealed by comparative genomic analyses. Peptides, 27 (12):3154-3164.

4.     Cockcroft JR, Noon JP, Gardner-Medwin J, Bennett T (1997). Haemodynamic effects of adrenomedullin in human resistance and capacitance vessels. Br J Clin Pharmacol, 44(1):57-60.

5.     Shimekake Y, Nagata K, Ohta S, Kambayashi Y, Teraoka H, Kitamura K, Eto T, Kangawa K, Matsuo H (1995). Adrenomedullin stimulates two signal transduction pathways, cAMP accumulation and Ca2+ mobilization, in bovine aortic endothelial cells. J Biol Chem, 270: 4412-4417.

6.     Yanagawa B, Nagaya N (2007). Adrenomedullin: molecular mechanisms and its role in cardiac disease. Amino Acids, 32 (1):157-164.

7.     Vesely DL (2003). Natriuretic peptides and acute renal failure. Am J Physiol Renal Physiol, 285 (2):167-177.

8.     Ruzicska E, Toth M, Tulassay Z, Somogyi A (2001). Adrenomedullin and diabetes mellitus. Diabetes Metab Res Rev, 17 (5):321-329.