RNA RECOGNITION BY LA AUTOANTIGEN

这个子项目是利用由NIH/NCRR资助的中心拨款提供的资源的许多研究子项目之一。子项目和调查员(PI)可能从另一个NIH来源获得了主要资金，因此可能会出现在其他CRISE条目中。列出的机构是针对中心的，而不一定是针对调查员的机构。我们的研究工作集中在人La NTD蛋白-RNA复合体的结构上，以确定以新生RNA转录物的3‘端为靶点的La自身抗原结合亲和力和特异性的分子基础。我们用核磁共振波谱质量来筛选一系列以UUUOH3‘-末端为末端的RNA低聚物，以考察它们在溶液中与La NTD形成络合物的能力。发现La NTD与UUUOH3-mer和9-mer RNA低聚体形成的3‘-UUUOH端的配合物具有最好的核磁共振谱，产生的衍射性晶体质量较高。晶体属于空间群P21212，每个不对称单元有两个络合物分子，在国产X射线源上衍射率为2.9A。利用APS 19ID光束线天然晶体的X射线数据，用分子置换方法找到了部分结构溶液。我们结晶了硒蛋氨酸(SeMet)蛋白的络合物，得到了用于结构测定的实验相。我们的长期目标是研究单个NTD和CTD以及全长La蛋白识别RNA的基础。

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. Our research efforts are focused on the structure of human La NTD protein-RNA complex to define the molecular basis for binding affinity and specificity of La autoantigen targeted to the 3¿¿¿-end of nascent RNA transcripts. We have used NMR spectral quality to screen a series of RNA oligomers terminating with UUUOH 3¿¿¿-ends for their ability to form complexes with La NTD in solution. The best NMR spectra were found for complexes of La NTD with the UUUOH 3-mer and 9-mer RNA oligomers containing 3¿¿¿-UUUOH ends, with the later yielding diffraction quality crystals. The crystals belong to space group P21212 with two complex molecules per asymmetric unit and diffracted to 2.9 A at home X-ray source. Partial structure solution has been found by molecular replacement method using x-ray data collected from native crystal at 19ID beamline at APS. We have crystallized complex with selenomethionine (SeMet) protein to obtain experimental phases for structure determination. Our long-term goal is to investigate basis of RNA recognition by the individual NTD and CTD as well as full-length La protein.