MAD EXPERIMENT OF GR DICER PROTEIN CRYSTAL; HIGH RESOLUTION DATA COLLECTION OF E

本子项目是利用由NIH/NCRR资助的中心赠款提供的资源的众多研究子项目之一。子项目和研究者（PI）可能已经从另一个NIH来源获得了主要资金，因此可以在其他CRISP条目中表示。列出的机构是中心的，不一定是研究者的机构。我们最近获得了Dicer的晶体，Dicer是RNA干扰（RNAi）途径中的一种关键酶，它可以衍射x射线到原子分辨率。这种大型二聚体蛋白的分子结构对于理解RNA如何在体内被识别和切割成特定长度的产物是至关重要的，这是通过靶向mRNA降解或翻译阻滞来干扰RNA的第一步。据我们所知，这是RNAi途径中第一个高质量晶体可用的酶，其结构可能对我们对RNAi分子机制的理解产生重大影响。硒代蛋氨酸结合晶体目前已经准备好在MAD实验中使用。真细菌信号识别粒子（SRP）由于其有限的复杂性，是高分辨率结构研究的一个很好的模型，以解决其功能。我最近获得了功能性大肠杆菌SRP和SRP受体（SR）复合物的晶体，并收集了高达8 A分辨率的初步衍射数据。一个高分辨率的结构模型，连同随后的生化实验，将提供SRP的重要功能，包括如何识别信号肽，以及SRP和SR的GTPase活性如何在不同的功能状态下被调节。

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. We have recently obtained crystals of Dicer, a key enzyme in the RNA interference (RNAi) pathway, that diffract X-rays to atomic resolution. The molecular structure of this large dimeric protein will be central to understanding how RNA is recognized and cleaved into specific length products in vivo as the first step in RNA interference by targeted mRNA degradation or translational arrest. To our knowledge this is the first enzyme in the RNAi pathway for which high-quality crystals are available, and the structure is likely to have a major impact on our understanding of the molecular mechanism of RNAi. Selenomethionine incorporated crystals are currently in hand and ready for use in MAD experiments. The eubacterial Signal Recognition Particle (SRP), because of its limited complexity, is an excellent model for high-resolution structural studies to address its function. I have recently obtained crystals of the functional E.Coli SRP and SRP receptor (SR) complex, and collected preliminary diffraction data up to 8 A resoltion. A high-resolution structure model, together with subsequent biochemical experiments, should provide insights into the important functions of SRP including how the signal peptide is recognized, and how the GTPase activities of SRP and SR are regulated at different functional states.