ID OF ENDOGENOUS LIGAND FOR DIOXAN ACTIVATED RECEPTOR (AH RECEPTOR)

二恶烷激活受体（AH 受体）内源配体的 ID

• 批准号: 7420539
• 负责人: Hector DeLuca
• 金额: $ 0.89万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-04-20 至 2007-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7420539
• 关键词: gel; ligands; receptor; reduction; reporter genes; role

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Dioxin is a major environmental toxin that binds to the Ah receptor activating a number of genes, resulting in toxicity and a number of changes in the reproductive system of fish. The biological role of the Ah receptor under normal conditions remains unknown. Using an Ah receptor-responsive reporter gene system and a selective extraction technique, we have successfully solubilized the endogenous ligand for this receptor from porcine lung. We have successfully purified this ligand using batch silica gel absorption and elution, large scale silica gel chromatography, followed by three high performance liquid chromatographic steps. This produces a homogenous substance that retains transcriptional activity in the reporter gene system. We have both low and high resolution mass spectra on this ligand as well as an ultraviolet absorption spectra. The high resolution mass spectra has given us an empirical formula which is C18H12N2O4. Borohydride reduction of the mole cule destr oys its biological activity and its chromophore. So far we have not determined the molecular weight of the reduction product, but in any case even with this information we are unable to identify positively the structure of the ligand. It is very clear that nuclear magnetic resonance spectroscopy would provide us the missing information, allowing identification of this much sought-after substance. We believe NMR on a 1 microgram sample in the 500 MHz instrument would provide us the missing information. If this fails, we believe that a short run on the 750 MHz instrument would certainly provide the resolution we need. Once identification has been achieved, we will immediately begin chemical synthesis of the ligand and begin to determine its biological role.

该子项目是利用NIH/NCRR资助的中心赠款提供的资源的许多研究子项目之一。子项目和研究者（PI）可能从另一个NIH来源获得主要资金，因此可以在其他CRISP条目中表示。所列机构为中心，不一定是研究者所在机构。二恶英是一种主要的环境毒素，它与Ah受体结合，激活一些基因，导致毒性和鱼类生殖系统的一些变化。Ah受体在正常条件下的生物学作用仍然未知。使用Ah受体响应报告基因系统和选择性提取技术，我们已经成功地溶解了来自猪肺的该受体的内源性配体。我们已经成功地纯化了这种配体使用批量硅胶吸附和洗脱，大规模硅胶色谱，然后通过三个高效液相色谱步骤。这产生了在报告基因系统中保留转录活性的同质物质。我们有这个配体的低分辨率和高分辨率质谱以及紫外吸收光谱。高分辨质谱给出了经验分子式C18 H12 N2 O 4。分子的硼氢化物还原破坏了其生物活性和发色团。到目前为止，我们还没有确定还原产物的分子量，但无论如何，即使有了这些信息，我们也无法确定配体的结构。很明显，核磁共振光谱将为我们提供缺失的信息，从而可以识别这种备受追捧的物质。我们相信，在500 MHz仪器中对1微克样品进行NMR将为我们提供缺失的信息。如果失败，我们相信在750 MHz仪器上进行短期运行肯定会提供我们需要的分辨率。一旦鉴定完成，我们将立即开始化学合成配体，并开始确定其生物学作用。