Dobbyn Helen C, McEwan Paul A, Krause Andre, Novak-Frazer Lily, Bella Jordi, O'Keefe Raymond T
School of Pharmacy, Centre for Biomolecular Sciences, University of Nottingham, Nottingham NG7 2RD, UK.
Biochem Biophys Res Commun. 2007 Sep 7;360(4):857-62. doi: 10.1016/j.bbrc.2007.06.163. Epub 2007 Jul 9.
Snu13p is a Saccharomyces cerevisiae protein essential for pre-messenger RNA splicing and pre-ribosomal RNA processing. Snu13p binds U4 snRNA of the spliceosome and box C/D snoRNAs of the pre-ribosomal RNA processing machinery to induce assembly of each ribonucleoprotein complex. Here, we present structural and biochemical analysis of Snu13p. The crystal structure of Snu13p reveals a region of the protein which could be important for protein interaction during ribonucleoprotein assembly. Using the structure of Snu13p we have designed the first temperature-sensitive mutants in Snu13p, L67W and I102A. Wild-type and mutant Snu13p proteins were assayed for binding to U4 snRNA and U3 snoRNA. Both temperature-sensitive mutants displayed significantly reduced RNA binding compared to wild-type protein. As the temperature-sensitive mutations are not in the known RNA binding region of Snu13p this indicates that these mutants indirectly influence the RNA binding properties of Snu13p. This work provides insight into Snu13p function during ribonucleoprotein assembly.
Snu13p是一种酿酒酵母蛋白，对于前体信使RNA剪接和前体核糖体RNA加工至关重要。Snu13p与剪接体的U4 snRNA以及前体核糖体RNA加工机制的C/D盒小核仁RNA结合，以诱导每个核糖核蛋白复合体的组装。在此，我们展示了对Snu13p的结构和生化分析。Snu13p的晶体结构揭示了该蛋白的一个区域，该区域在核糖核蛋白组装过程中对于蛋白质相互作用可能很重要。利用Snu13p的结构，我们设计了Snu13p中的首个温度敏感突变体L67W和I102A。对野生型和突变型Snu13p蛋白进行了与U4 snRNA和U3 snoRNA结合的检测。与野生型蛋白相比，这两个温度敏感突变体均显示出显著降低的RNA结合能力。由于温度敏感突变并不在Snu13p已知的RNA结合区域，这表明这些突变体间接影响了Snu13p的RNA结合特性。这项工作为深入了解Snu13p在核糖核蛋白组装过程中的功能提供了依据。
