DAz-2
(Synonyms: DCP-N3) 目录号 : GC43382A chemical probe for sulfenic acid detection
Cas No.:1176905-54-6
Sample solution is provided at 25 µL, 10mM.
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Quality Control & SDS
- View current batch:
- Purity: >98.00%
- COA (Certificate Of Analysis)
- SDS (Safety Data Sheet)
- Datasheet
DAz-2 is a cell-permeable chemical probe used to detect cysteine oxidation in proteins. Redox-sensitive cysteine residues in proteins may function as sensors of reactive oxygen species (ROS) and also serve as molecular switches, activating or deactivating proteins, following a change in oxidation state. Modification of protein function through the reversible oxidation of cysteine is emerging as a biologically relevant signal transduction mechanism. Sulfenic acid is the initial oxidation product of cysteine by relatively mild oxidizing agents such as hydrogen peroxide. Sulfenic acid can be reduced back to the free thiol or be further oxidized to sulfinic and sulfonic acids.[1] DAz-2 is a cell-permeable chemical probe that reacts specifically with sulfenic acid-modified proteins.[2] The azido group of DAz-2 provides a method for selective conjugation to phosphine- or alkynyl- derivatized reagents, such as biotin or various fluorophores, for subsequent analysis of the labeled proteins. Use of DAz-2 in HeLa cells followed by Staudinger ligation to biotin and subsequent LC-MS/MS analysis, led to the identification of 193 sulfenic acid-modified proteins having a diverse range of functions.[2]
Reference:
[1]. Reddie, K.G., and Carroll, K.S. Expanding the functional diversity of proteins through cysteine oxidation. Current Opinion in Chemical Biology 12(6), 746-754 (2008).
[2]. Leonard, S.E., Reddie, K.G., and Carroll, K.S. Mining the thiol proteome for sulfenic acid modifications reveals new targets for oxidation in cells. ACS Chem. Biol. 4(9), 783-799 (2009).
| Cas No. | 1176905-54-6 | SDF | |
| 别名 | DCP-N3 | ||
| 化学名 | 4-(3-azidopropyl)cyclohexane-1,3-dione | ||
| Canonical SMILES | O=C(C1)C(CCCN=[N+]=[N-])CCC1=O | ||
| 分子式 | C9H13N3O2 | 分子量 | 195.2 |
| 溶解度 | 14 mg/ml in DMSO, 20 mg/ml in DMF, 25 mg/ml in Ethanol | 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 | |||
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1 mg | 5 mg | 10 mg |
| 1 mM | 5.123 mL | 25.6148 mL | 51.2295 mL |
| 5 mM | 1.0246 mL | 5.123 mL | 10.2459 mL |
| 10 mM | 0.5123 mL | 2.5615 mL | 5.123 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。
Redox regulation of RAD51 Cys319 and homologous recombination by peroxiredoxin 1
Redox Biol 2022 Oct;56:102443.PMID:36058112DOI:10.1016/j.redox.2022.102443.
RAD51 is a critical recombinase that functions in concert with auxiliary mediator proteins to direct the homologous recombination (HR) DNA repair pathway. We show that Cys319 RAD51 possesses nucleophilic characteristics and is important for irradiation-induced RAD51 foci formation and resistance to inhibitors of poly (ADP-ribose) polymerase (PARP). We have previously identified that cysteine (Cys) oxidation of proteins can be important for activity and modulated via binding to peroxiredoxin 1 (PRDX1). PRDX1 reduces peroxides and coordinates the signaling actions of protein binding partners. Loss of PRDX1 inhibits irradiation-induced RAD51 foci formation and represses HR DNA repair. PRDX1-deficient human breast cancer cells and mouse embryonic fibroblasts display disrupted RAD51 foci formation and decreased HR, resulting in increased DNA damage and sensitization of cells to irradiation. Following irradiation cells deficient in PRDX1 had increased incorporation of the sulfenylation probe DAz-2 in RAD51 Cys319, a functionally-significant, thiol that PRDX1 is critical for maintaining in a reduced state. Molecular dynamics (MD) simulations of dT-DNA bound to a non-oxidized RAD51 protein showed tight binding throughout the simulation, while dT-DNA dissociated from an oxidized Cys319 RAD51 filament. These novel data establish RAD51 Cys319 as a functionally-significant site for the redox regulation of HR and cellular responses to IR.
Activity of the tetrapyrrole regulator CrtJ is controlled by oxidation of a redox active cysteine located in the DNA binding domain
Mol Microbiol 2012 Aug;85(4):734-46.PMID:22715852DOI:10.1111/j.1365-2958.2012.08135.x.
CrtJ from Rhodobacter capsulatus is a regulator of genes involved in the biosynthesis of haem, bacteriochlorophyll, carotenoids as well as structural proteins of the light harvesting-II complex. Fluorescence anisotropy-based DNA-binding analysis demonstrates that oxidized CrtJ exhibits ~20-fold increase in binding affinity over that of reduced CrtJ. Liquid chromatography electrospray tandem ionization mass spectrometric analysis using DAz-2, a sulfenic acid (-SOH)-specific probe, demonstrates that exposure of CrtJ to oxygen or to hydrogen peroxide leads to significant accumulation of a sulfenic acid derivative of Cys420 which is located in the helix-turn-helix (HTH) motif. In vivo labelling with 4-(3-azidopropyl)cyclohexane-1,3-dione (DAz-2) shows that Cys420 also forms a sulfenic acid modification in vivo when cells are exposed to oxygen. Moreover, a Cys420 to Ala mutation leads to a ~60-fold reduction of DNA binding activity while a Cys to Ser substitution at position 420 that mimics a cysteine sulfenic acid results in a ~4-fold increase in DNA binding activity. These results provide the first example where sulfenic acid oxidation of a cysteine in a HTH-motif leads to differential effects on gene expression.
Mining the thiol proteome for sulfenic acid modifications reveals new targets for oxidation in cells
ACS Chem Biol 2009 Sep 18;4(9):783-99.PMID:19645509DOI:10.1021/cb900105q.
Oxidation of cysteine to sulfenic acid has emerged as a biologically relevant post-translational modification with particular importance in redox-mediated signal transduction; however, the identity of modified proteins remains largely unknown. We recently reported DAz-1, a cell-permeable chemical probe capable of detecting sulfenic acid modified proteins directly in living cells. Here we describe DAz-2, an analogue of DAz-1 that exhibits significantly improved potency in vitro and in cells. Application of this new probe for global analysis of the sulfenome in a tumor cell line identifies most known sulfenic acid modified proteins: 14 in total, plus more than 175 new candidates, with further testing confirming oxidation in several candidates. The newly identified proteins have roles in signal transduction, DNA repair, metabolism, protein synthesis, redox homeostasis, nuclear transport, vesicle trafficking, and ER quality control. Cross-comparison of these results with those from disulfide, S-glutathionylation, and S-nitrosylation proteomes reveals moderate overlap, suggesting fundamental differences in the chemical and biological basis for target specificity. The combination of selective chemical enrichment and live-cell compatibility makes DAz-2 a powerful new tool with the potential to reveal new regulatory mechanisms in signaling pathways and identify new therapeutic targets.
Inactivation of thiol-dependent enzymes by hypothiocyanous acid: role of sulfenyl thiocyanate and sulfenic acid intermediates
Free Radic Biol Med 2012 Mar 15;52(6):1075-85.PMID:22248862DOI:10.1016/j.freeradbiomed.2011.12.024.
Myeloperoxidase (MPO) forms reactive oxidants including hypochlorous and hypothiocyanous acids (HOCl and HOSCN) under inflammatory conditions. HOCl causes extensive tissue damage and plays a role in the progression of many inflammatory-based diseases. Although HOSCN is a major MPO oxidant, particularly in smokers, who have elevated plasma thiocyanate, the role of this oxidant in disease is poorly characterized. HOSCN induces cellular damage by targeting thiols. However, the specific targets and mechanisms involved in this process are not well defined. We show that exposure of macrophages to HOSCN results in the inactivation of intracellular enzymes, including creatine kinase (CK) and glyceraldehyde-3-phosphate dehydrogenase (GAPDH). In each case, the active-site thiol residue is particularly sensitive to oxidation, with evidence for reversible inactivation and the formation of sulfenyl thiocyanate and sulfenic acid intermediates, on treatment with HOSCN (less than fivefold molar excess). Experiments with DAz-2, a cell-permeable chemical trap for sulfenic acids, demonstrate that these intermediates are formed on many cellular proteins, including GAPDH and CK, in macrophages exposed to HOSCN. This is the first direct evidence for the formation of protein sulfenic acids in HOSCN-treated cells and highlights the potential of this oxidant to perturb redox signaling processes.
Isotope-coded chemical reporter and acid-cleavable affinity reagents for monitoring protein sulfenic acids
Bioorg Med Chem Lett 2011 Sep 1;21(17):5015-20.PMID:21601453DOI:10.1016/j.bmcl.2011.04.115.
We have developed an approach that allows relative quantification of protein sulfenic acids using a pair of light and heavy isotope labled probes, DAz-2 and d(6)-DAz-2. In conjunction with a new complementary acid-cleavable linker, Yn-ACL, we demonstrate that tagged peptides are successfully labeled, enriched, and fully characterized by LC-MS/MS analysis. Overall, this method can be applied to map sites of cysteine oxidation and compare protein sulfenylation in normal and disease states.