3-(3-氨基-3-羧丙基)尿苷(acp3U)是细菌和真核生物tRNA核心区广泛存在的保守修饰，在肿瘤细胞中水平增高，是潜在的体内肿瘤标记物。TapT是近期鉴定的新型tRNA acp3U修饰酶，负责tRNA 可变环区47位的acp3U修饰。TapT属于SAM依赖修饰酶，序列比对显示TapT的底物识别和酶活催化机制和现有acp修饰酶存在差异。本项目旨在通过X-射线衍射晶体学等结构生物学的方法解析TapT自由态、SAM结合态、目标RNA结合态的三维结构；基于结构，结合定点突变、等温量热滴定、体外酶活实验等生物化学和分子生物学方法，探讨TapT的底物识别机制，确定其酶活催化机制；并以结构为出发点进一步比较分析它与Tsr3的底物催化机制的异同点，比较分析两种重要核酸分子tRNA与rRNA acp3U修饰发生的异同点，进而推动其在生物化学和分子生物学以及肿瘤发生学等领域的应用。

3-(3-amino-3-carboxypropyl)uridine (acp3U) is a highly conserved modification found in variable- and D-loops of tRNAs in Bacteria and Eukarya, which is also a potential tumor marker in vivo due to its increased level in tumor cells. TapT is a recently identified enzyme responsible for acp3U modification at position 47 of E.coli tRNAs in the presence of S-adenosylmethionine (SAM). Sequence alignments reveal that TapT is diverse from other acp modification enzymes, indicating that TapT adopts specific substrate recognition mode and catalytic mechanism. However, the specific substrate binding details and catalytic mechanism of TapT remains unknown with the absence of TapT structures. This project aims to figure out the three-dimensional structures of TapT with free state, SAM bound state or target RNA bound state. Based on the structure and related functional experiments, we will try to discover the substrate recognition mode and the enzyme catalytic mechanism. Furthermore, with structure and sequence comparing, we will try to discuss the similarities and differences of the substrate catalytic mechanism between TapT and Tsr3, which could finally promote its application in biochemistry, molecular biology and tumor diagnostics.