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118850-72-9,Anti-Inflammatory Peptide 2,H2N-His-Asp-Met-Asn-Lys-Val-Leu-Asp-Leu-COOH,H2N-HDMNKVLDL-OH,杭州专肽生物的产品

Anti-Inflammatory Peptide 2

HDMNKVLDL。

编号:125848

CAS号:118850-72-9

单字母:H2N-HDMNKVLDL-OH

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  • 编号:125848
    中文名称:Anti-Inflammatory Peptide 2
    英文名:Anti-Inflammatory Peptide 2
    CAS号:118850-72-9
    单字母:H2N-HDMNKVLDL-OH
    三字母:H2N

    N端氨基

    -His

    组氨酸

    -Asp

    天冬氨酸

    -Met

    甲硫氨酸

    -Asn

    天冬酰胺

    -Lys

    赖氨酸

    -Val

    缬氨酸

    -Leu

    亮氨酸

    -Asp

    天冬氨酸

    -Leu

    亮氨酸

    -OH

    C端羧基

    氨基酸个数:9
    分子式:C46H77N13O15S1
    平均分子量:1084.25
    精确分子量:1083.54
    等电点(PI):7.55
    pH=7.0时的净电荷数:0.21
    平均亲水性:0.25555555555556
    疏水性值:-0.43
    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
    生成周期:2-3周
    储存条件:负80℃至负20℃
    标签:抗炎肽(Anti-Inflammatory Peptide)    炎症研究   

  • HDMNKVLDL.

    Definition
    Small, anti-inflammatory peptides are useful to inhibit inflammation of a mammal's skin, mucous membranes, or lacerations of the musculature or injury to the brain or leakage of fluids into the air spaces of the lungs.

    Discovery
    Camussi G in 1986 described platelet-activating factor (PAF) as a phospholipid (1-0-alkyl-2-sn-acetyl-glycero-3-phosphocholine) mediator of inflammation and endotoxic shock. Polymorphonuclear neutrophils (PMN), peritoneal macrophages, vascular endothelial cells, basophils, and platelets synthesize PAF rapidly after appropriate stimuli 1.  For example, TNF or phagocytosis promotes synthesis and release of PAF in PMN or macrophages within 10 min 2.  These stimuli induce phospholipase A2 (PLA2) activity that cleaves membrane phospholipids into lyso-PAF and arachidonic acid. Thrombin stimulates PAFsynthesis in endothelial cells also within 10 min 3.  Other proteinases, such as elastase, stimulate PAF synthesis rapidly in PMN, macrophages, and endothelial cells, and induce PLA2 and acetyltransferase activity. Conversely, different proteinase inhibitors block PAF synthesis induced by TNF. These findings led to the hypothesis that proteinases added to cells and cellular proteinases activated by TNF cleave proteins inhibitory for PLA2, such as lipocortins 4.  Lipocortins belong to a family of related proteins that mediate the anti-inflammatory activity of corticosteroids.

    Structural Characteristics
    Cloning and sequencing of lipocortins cDNA has provided the amino acid sequence of anti-inflammatory proteins. Steroid induced protein with PLA2 inhibitory activity is uteroglobin, a rabbit secretory protein 5.  Two identical subunits of 70 amino acids form uteroglobin; lipocortin I and II comprise four nonidentical repeats of 70 amino acids. Miele et al., have noticed a striking sequence similarity between amino acid residues 40-46 of uteroglobin and 247-253 of lipocortin I, repeat 3. Synthetic peptides designated "antiflammins" that correspond to such sequences show potent PLA2 inhibitory activity in vitro and an antiinflammatory effect on carrageenan induced rat foot pad edema in vivo 6, 7. SV-IV is a basic, thermostable, secretory protein of low Mr (9758) that is synthesized by rat seminal vesicle (SV) epithelium under strict androgen transcriptional control. By using two different experimental, data were obtained showing that in this protein: (a) the immunomodulatory activity is related to the structural integrity of the whole molecule; (b) the anti-inflammatory activity is located in the N-terminal region of the molecule, the 8-16 peptide fragment being the most active; (c) the identified anti-inflammatory peptide derivatives do not seem to possess pro-coagulant activity, even though this particular function has been located in the 1-70 segment of the molecule 8. The peptide HDMNKVLDL (antiflammin-2) inhibits the synthesis of PAF induced by TNF or phagocytosis in rat macrophages and human neutrophils, and by thrombin in vascular endothelial cells. The peptide MQMKKVLDS (antiflammin-1) is less inhibitory than antiflammin-2 for macrophages and not inhibitory for neutrophils after a 5-min preincubation 9.

    Mode of Action
    PLA2 are a family of esterases that initiate the arachidonic acid cascade, resulting in the production of numerous inflammatory mediators. Lipocortins are inhibitors of PLA2. Peptides corresponding to a region of high amino-acid sequence similarity between uteroglobin and lipocortin I have potent PLA2 inhibitory activity. Several synthetic peptides corresponding to the region of highest similarity were designed by Miele, L. et al., The most effective anti-inflammatory nonapeptides, termed antiflammins (AFs) correspond to uteroglobin residues 39–47 and lipocortin-1 residues 246–254. Both peptides are PLA2 inhibitors in-vitro and are effective in a classic model of acute inflammation in carrageenan-induced rat footpad edema. The nonapeptides have anti-inflammatory effects in-vitro and in-vivo 10. Antiflammin-1 is inactivated by neutrophils secretory products, possibly oxidizing agents. Synthesis of PAF is inhibited by antiflammin-2 without an appreciable lag, but this inhibition is reversed when neutrophils or macrophages are washed and incubated in fresh medium. Antiflammins must be continuously present to inhibit PAF synthesis. Antiflammins block activation of the acetyltransferase required for PAF synthesis, suggesting that this enzyme is another target for the inhibitory activity of antiflammins. These peptides inhibit neutrophil aggregation and chemotaxis induced by complement component C5a 9.

    Functions

    Bee venom, in a study of phospholipase-A2,3 nearly 1 kg of freeze-dried bee venom was fractionated and tested all the fractions for anti-inflammatory activity. Experiments showed activity to be associated with the basic peptide fraction 11. 

    PAF synthesis, anti-inflammatory peptides inhibit PAF synthesis in PMN, macrophages, and endothelial cells stimulated by TNF, phagocytosis, or proteinases 9.

    PMN aggregation and chemotaxis, the antiflammins inhibit also PMN aggregation and chemotaxis, and suppress the inflammatory reaction induced in rat skin by in situ formation of immune complexes or by intradermal injection of TNF and complement component C5a 9.

    References

    1.     Camussi G (1986). Potential role of platelet-activating factor in renal pathophysiology. Kidney Int., 29: 469-477.
    2.      Camussi G, Bussolino F, Salvidio G, Baglioni C (1987). Tumor necrosis factor/cachectin stimulates rat peritoneal macrophages and human endothelial cells to synthesize and release platelet activating factor. J. Exp. Med., 166:1390-1404.
    3.     Prescott SM (1984). Human endothelial cells in culture produce platelet-activating factor (1-alkyl-2-acetyl-sn-glyceryl-3-phosphocholine) when stimulated with thrombin. PNAS., 81:3534-3538.
    4.     Camussi G, Tetta C, Bussolino F, Baglioni C (1988). Synthesis and release of platelet-activating factor is inhibited by plasma a,proteinase inhibitor or citantichymotrypsin and stimulated by proteinases . J. Exp. Med., 168:1293-1306.
    5.     Levin SW, Butler JD, Schumacher UK, Wightman PD, Mukherjee AB (1986). Uteroglobin inhibits phospholipase A2 activity. Life Sci., 38:1813-1819.
    6.     Morize I, Surcouf E, Vaney MC, Epelboin Y, Buehner M, Fridlansky F, Milgrom E, Mornon JP (1987) . Refinement of the C222 crystal formofoxidized uteroglobin at 1 .34 A resolution. J. Mol. Biol., 194:725-739.
    7.   Miele L, Cordella-Miele E, Facchiano A, Mukherjee AB (1988). Novel anti-inflammatory peptides from the region of highest similarity between uteroglobin and lipocortin I. Nature, 335:726-730.
    8.     Ialenti A, Santagada V, Caliendo G, Severino B, Fiorino F, Maffia P, Ianaro A, Morelli F, Di Micco B, Cartenì M, Stiuso P, Metafora V, Metafora S (2001).  Synthesis of novel anti-inflammatory peptides derived from the amino-acid sequence of the bioactive protein SV-IV. FEBS Journal, 268(12):3399-3406.
    9.     Camussi G, Tetta C, Bussolino F, Baglioni C (1990). Antiinflamatory peptides (antiinflamins) inhibit synthesis of platelet-activating factor, neutrophil aggregation and chemotaxis, and interdermal inflammatory reactions. J. Exp. Med., 171:913-927.
    10.  Sohn J, Kim TI, Yoon YH, Kim JY, Kim SY (2003). Novel transglutaminase inhibitors reverse the inflammation of allergic conjunctivitis. J. Clin. Invest., 111(1):121-128.
    11.   Billingham ME, Morley J, Hanson JM, Shipolini RA, Vernon CA (1973). Letter: An anti-inflammatory peptide from bee venom.  Nature, 245(5421):163-164.

  • DOI名称
    10.1021/ac048282zSecondary reactions and strategies to improve quantitative protein footprinting下载
    10.1016/j.intimp.2005.08.001Effects of antiflammins on transglutaminase and phospholipase A2 activation by transglutaminase下载
  • 多肽H2N-His-Asp-Met-Asn-Lys-Val-Leu-Asp-Leu-COOH的合成步骤:

    1、合成CTC树脂:称取2.79g CTC Resin(如初始取代度约为0.74mmol/g)和2.48mmol Fmoc-Leu-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6.19mmol 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-Leu-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-Leu-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6.19mmol Fmoc-Asp(OtBu)-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12.39mmol DIPEA,5.88mmol 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-Asp(OtBu)-Leu-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-Asp(OtBu)-Leu-CTC Resin

    Fmoc-Leu-Asp(OtBu)-Leu-CTC Resin

    H2N-Leu-Asp(OtBu)-Leu-CTC Resin

    Fmoc-Val-Leu-Asp(OtBu)-Leu-CTC Resin

    H2N-Val-Leu-Asp(OtBu)-Leu-CTC Resin

    Fmoc-Lys(Boc)-Val-Leu-Asp(OtBu)-Leu-CTC Resin

    H2N-Lys(Boc)-Val-Leu-Asp(OtBu)-Leu-CTC Resin

    Fmoc-Asn(Trt)-Lys(Boc)-Val-Leu-Asp(OtBu)-Leu-CTC Resin

    H2N-Asn(Trt)-Lys(Boc)-Val-Leu-Asp(OtBu)-Leu-CTC Resin

    Fmoc-Met-Asn(Trt)-Lys(Boc)-Val-Leu-Asp(OtBu)-Leu-CTC Resin

    H2N-Met-Asn(Trt)-Lys(Boc)-Val-Leu-Asp(OtBu)-Leu-CTC Resin

    Fmoc-Asp(OtBu)-Met-Asn(Trt)-Lys(Boc)-Val-Leu-Asp(OtBu)-Leu-CTC Resin

    H2N-Asp(OtBu)-Met-Asn(Trt)-Lys(Boc)-Val-Leu-Asp(OtBu)-Leu-CTC Resin

    Fmoc-His(Trt)-Asp(OtBu)-Met-Asn(Trt)-Lys(Boc)-Val-Leu-Asp(OtBu)-Leu-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-His(Trt)-Asp(OtBu)-Met-Asn(Trt)-Lys(Boc)-Val-Leu-Asp(OtBu)-Leu-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-His-Asp-Met-Asn-Lys-Val-Leu-Asp-Leu-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:

    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

    以上所有内容,为专肽生物原创内容,请勿发布到其他网站上。

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