CATALYTIC DOMAIN OF CHICKEN TRYPTOPHAN HYDROXYLASE ISOFORM 1

鸡色氨酸羟化酶异构体 1 的催化结构域

• 批准号: 8363390
• 负责人: HANS ERIK CHRISTENSEN
• 金额: $ 0.29万
• 依托单位: BROOKHAVEN SCIENCE ASSOC-BROOKHAVEN LAB
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-07-01 至 2012-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8363390
• 关键词: Anabolism; Aromatic Amino Acids; Binding; Biochemistry; Brain; Catalytic Domain; Chickens; Crystallization; Crystallography; Data; Disease; Eating Disorders; Enzymes; Funding; Grant; Iron; Knowledge; Laboratories; Ligands; Light; Mental Depression; Methods; Mixed Function Oxygenases; Molecular; National Center for Research Resources; Neurotransmitters; Obsessive-Compulsive Disorder; Principal Investigator; Process; Protein Isoforms; Publishing; Research; Research Infrastructure; Resolution; Resources; Schizophrenia; Serotonin; Shapes; Solutions; Source; Structure; Structure-Activity Relationship; Synchrotrons; Time; Tryptophan 5-monooxygenase; United States National Institutes of Health; Universities; Variant; cofactor; cost; neuropsychiatry

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Depression, eating disorders, schizophrenia, OCD (obsessive compulsive disorder) and many other neuropsychiatric disorders are presumably caused by imbalance in the level of the neurotransmitter serotonin in the brain.The iron-containing enzyme tryptophan hydroxylase (TPH) catalyzes the rate-limiting step in the biosynthesis of serotonin and knowledge on the structure and function of this enzyme is thus important for the molecular understanding of the disorders mentioned above. The complete mechanism of TPH is not known.TPH is known to be notoriously unstable and difficult to purify, and only 5 crystal structures of the double truncated enzyme have been published. Our group has developed methods to purify stable solutions of several TPH variants, which have now been crystallized. One structure has already been solved and was published in Biochemistry in 2008 (PDB ID 3E2T). This was the first structure of any aromatic amino acid hydroxylase with the natural substrate bound. Now we have crystals of this same variant of TPH, with only substrate (the previous structure also contained an unnatural ligand from the crystallization solution) and with substrate and cofactor bound. Determination of these structures will allow us to investigate the structural changes which occur upon substrate and cofactor binding. Together with CD/MCD data collected at The Solomon Laboratory at Stanford University, the structures will give a complete picture of the structure/function relationship in TPH1. The crystals of TPH1 are between 40 and 100 um, clearly three dimensional and nicely shaped with smooth edges. The last crystal diffracted to a resolution of 1.90 ¿, however these new crystals appeared after shorter time (3 days vs. 6 months) so the actual resolution is not known.As we have several crystals for each of the two possible structures, a scouting process for the-best-in-the-lot using automounters would be preferable. The structures will be solved with molecular replacement using the previous structure (3E2T).

这个子项目是许多利用资源的研究子项目之一 由NIH/NCRR资助的中心拨款提供。子项目的主要支持 而子项目的主要调查员可能是由其他来源提供的， 包括其它NIH来源。 列出的子项目总成本可能 代表子项目使用的中心基础设施的估计数量， 而不是由NCRR赠款提供给子项目或子项目工作人员的直接资金。 抑郁症，饮食失调，精神分裂，OCD强迫症和许多其他神经精神疾病可能是由大脑中神经递质血清素水平失衡引起的。含铁酶色氨酸羟化酶（TPH）催化-血清素是血清素生物合成中的一个限制步骤，了解这种酶的结构和功能对于从分子水平上了解这些疾病是很重要的上面提到的。TPH的完整机制尚不清楚，众所周知，TPH不稳定且难以纯化，仅发表了5种双截短酶的晶体结构。 我们的小组已经开发出纯化几种TPH变体的稳定溶液的方法，这些变体现在已经结晶。其中一个结构已经得到解决，并于2008年发表在《生物化学》（PDB ID 3E 2 T）上。这是芳香族氨基酸羟化酶与天然底物结合的第一个结构。现在我们有了TPH的这种相同变体的晶体，只有底物（以前的结构也包含来自结晶溶液的非天然配体），并且底物和辅因子结合。这些结构的测定将使我们能够研究底物和辅因子结合后发生的结构变化。结合斯坦福大学所罗门实验室收集的CD/MCD数据，这些结构将给出TPH 1结构/功能关系的完整图片。TPH 1的晶体在40和100 μ m之间，清晰的三维形状，边缘光滑。最后一个晶体衍射到1.90 <$的分辨率，然而这些新晶体出现后较短的时间（3天与6个月），所以实际的分辨率是未知的。因为我们有几个晶体的每一个两种可能的结构，侦察过程中最好的在该批次使用自动计数器将是可取的。将使用先前的结构（3E 2 T）通过分子置换解析结构。