Biotin-HPDP
(Synonyms: N-(6-[生物素胺]己基)-3-(2-吡啶二硫)丙酰胺,BiotinHPDP) 目录号 : GC11037
Biotin-HPDP 是一种巯基反应性生物素化试剂，可形成可逆的二硫键。
Cas No.:129179-83-5
Sample solution is provided at 25 µL, 10mM.
Biotin-HPDP is a sulfhydryl-reactive biotinylation reagent that forms a reversible disulfide linkage.Biotin-HPDP consists of a bicyclic Biotin ring structure, a 1, 6-diaminohexane attached to the side chain of biotin valonate, and a sulfhydryl reactive group at the end of the side chain[1]. It is used to label protein cysteines and other substrates that contain sulfhydryl groups.
After NEM alkylation, certain (but not all) residual free cysteine residues may react with biotin-HPDP, resulting in the coisolation of proteins containing such cysteine residues[2].
A kinetic study of the modification reactions that generate monothiophosphate disulfide linkages with either 5'-GMPS alone or 5'-GMPS-primed RNA as the substrate revealed that the second-order rate constants increased as the pH was decreased. For example, when the reaction pH was lowered from 8 to 4, the k2 value for the coupling reaction between N-(6-[biotinamido]hexyl)-3'-(2'-pyridyldithio)propionamide (biotin-HPDP) and GMPS increased 67-fold from 1.84 to 123 M(-1) x s(-1) [3].
References:
[1]: Bioconjugate Techniques , 2nd ed. By Greg T.Hermanson ?(Pierce Biotechnology, Thermo Fisher Scientific, Rockford, IL). ?Academic Press ?(an imprint of Elsevier): ?London, Amsterdam, Burlington, San Diego . 2008. ISBN 978-0-12-370501-3.
[2]: Zhou B, Wang Y, Yan Y, Mariscal J, Di Vizio D, Freeman MR, Yang W. Low-Background Acyl-Biotinyl Exchange Largely Eliminates the Coisolation of Non-S-Acylated Proteins and Enables Deep S-Acylproteomic Analysis. Anal Chem. 2019 Aug 6;91(15):9858-9866. doi: 10.1021/acs.analchem.9b01520. Epub 2019 Jul 11. PMID: 31251020; PMCID: PMC7451198.
[3]: Wu CW, Eder PS, Gopalan V, Behrman EJ. Kinetics of coupling reactions that generate monothiophosphate disulfides: implications for modification of RNAs. Bioconjug Chem. 2001 Nov-Dec;12(6):842-4. doi: 10.1021/bc0100612. PMID: 11716671.
Biotin-HPDP 是一种巯基反应性生物素化试剂，可形成可逆的二硫键。Biotin-HPDP 由双环生物素环结构、连接到生物素戊酸酯侧链的 1, 6-二氨基己烷和一个巯基反应性物质组成组在侧链的末端[1]。用于标记蛋白质半胱氨酸和其他含有巯基的底物。
NEM 烷基化后，某些（但不是全部）残留的游离半胱氨酸残基可能与生物素-HPDP 反应，导致含有此类半胱氨酸残基的蛋白质共分离[2]。
对以单独的 5'-GMPS 或以 5'-GMPS 引发的 RNA 作为底物产生单硫代磷酸盐二硫键的修饰反应的动力学研究表明，二级速率常数随着 pH 值的降低而增加。例如，当反应 pH 值从 8 降低到 4 时，N-(6-[生物素氨基]己基)-3'-(2'-吡啶二硫代)丙酰胺（生物素-HPDP）与 GMPS 之间的偶联反应的 k2 值从 1.84 增加 67 倍至 123 M(-1) x s(-1) [3]。
| Biotinylation method [1]: | |
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Sample |
S-nitrosylated proteins |
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Preparation method |
Soluble in DMSO or DMF. |
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Reaction Conditions |
50mM, 25 ℃ for 1 h |
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Applications |
Prepare biotin-HPDP as a 50mM suspension in DMSO, Dilute with DMF to a final concentration of 4 mM. Add 1:3 volume of Labeling Solution and 1:50 volume of Ascorbate Solution（50mM） to the blocked protein samples, incubate for 1 hour at 25°C. After that, add two volumes of −20°C acetone and incubate for 20 min at −20°C to remove the biotin-HPDP. At last, Add 15 µl of packed streptavidin-agarose per mg of protein used in the initial protein sample, to purify biotinylated proteins. Incubate the biotinylated proteins with the resin for 1 hour at room temperature. Wash the beads five times with 10 volumes of Neutralization Buffer + NaCl. Centrifuge at 200g for 5 s at room temperature between each wash. Incubate the beads with Elution Buffer to recover the bound proteins. To test for the protein of interest with specific antibodies in SDS-PAGE. |
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References: [1].Samie R. Jaffrey and Solomon H. Snyder. The Biotin Switch Method for the Detection of S-Nitrosylated Proteins. Science’s stke.2015. |
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| Cas No. | 129179-83-5 | SDF | |
| 别名 | N-(6-[生物素胺]己基)-3-(2-吡啶二硫)丙酰胺,BiotinHPDP | ||
| 化学名 | 5-[(3aS,4S,6aR)-2-oxo-1,3,3a,4,6,6a-hexahydrothieno[3,4-d]imidazol-4-yl]-N-[6-[3-(pyridin-2-yldisulfanyl)propanoylamino]hexyl]pentanamide | ||
| Canonical SMILES | C1C2C(C(S1)CCCCC(=O)NCCCCCCNC(=O)CCSSC3=CC=CC=N3)NC(=O)N2 | ||
| 分子式 | C24H37N5O3S3 | 分子量 | 539.78 |
| 溶解度 | 3mg/mL in DMSO | 储存条件 | Store at -20°C |
| General tips | 请根据产品在不同溶剂中的溶解度选择合适的溶剂配制储备液；一旦配成溶液，请分装保存，避免反复冻融造成的产品失效。 储备液的保存方式和期限：-80°C 储存时，请在 6 个月内使用，-20°C 储存时，请在 1 个月内使用。 为了提高溶解度，请将管子加热至37℃，然后在超声波浴中震荡一段时间。 |
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| Shipping Condition | 评估样品解决方案：配备蓝冰进行发货。所有其他可用尺寸：配备RT，或根据请求配备蓝冰。 | ||
| 制备储备液 | |||
 |
1 mg | 5 mg | 10 mg |
| 1 mM | 1.8526 mL | 9.263 mL | 18.5261 mL |
| 5 mM | 0.3705 mL | 1.8526 mL | 3.7052 mL |
| 10 mM | 0.1853 mL | 0.9263 mL | 1.8526 mL |
| 第一步：请输入基本实验信息（考虑到实验过程中的损耗，建议多配一只动物的药量） | ||||||||||
| 给药剂量 | mg/kg |  |
动物平均体重 | g | |
每只动物给药体积 | ul | |
动物数量 | 只 |
| 第二步：请输入动物体内配方组成（配方适用于不溶于水的药物；不同批次药物配方比例不同，请联系GLPBIO为您提供正确的澄清溶液配方） | ||||||||||
| % DMSO % % Tween 80 % saline | ||||||||||
| 计算重置 | ||||||||||
计算结果:
工作液浓度： mg/ml；
DMSO母液配制方法： mg 药物溶于 μL DMSO溶液（母液浓度 mg/mL，
体内配方配制方法：取 μL DMSO母液，加入 μL PEG300，混匀澄清后加入μL Tween 80，混匀澄清后加入 μL saline，混匀澄清。
1. 首先保证母液是澄清的；
2.
一定要按照顺序依次将溶剂加入，进行下一步操作之前必须保证上一步操作得到的是澄清的溶液，可采用涡旋、超声或水浴加热等物理方法助溶。
3. 以上所有助溶剂都可在 GlpBio 网站选购。
Quality Control & SDS
- View current batch:
- Purity: >98.00%
- COA (Certificate Of Analysis)
- SDS (Safety Data Sheet)
- Datasheet