RNA. 2014 Jan;20(1):24-35. doi: 10.1261/rna.039701.113. Epub 2013 Nov 15.
Recognition of the 5' splice site by group II introns involves pairing between an exon binding sequence (EBS) 1 within the ID3 stem-loop of domain 1 and a complementary sequence at the 3' end of exon 1 (IBS1). To identify the molecular basis for splice site definition of a group IIB ai5γ intron, we probed the solution structure of the ID3 stem-loop alone and upon binding of its IBS1 target by solution NMR. The ID3 stem was structured. The base of the ID3 loop was stacked but displayed a highly flexible EBS1 region. The flexibility of EBS1 appears to be a general feature of the ai5γ and the smaller Oceanobacillus iheyensis (O.i.) intron and may help in effective search of conformational space and prevent errors in splicing as a result of fortuitous base-pairing. Binding of IBS1 results in formation of a structured seven base pair duplex that terminates at the 5' splice site in spite of the potential for additional A-U and G•U pairs. Comparison of these data with conformational features of EBS1-IBS1 duplexes extracted from published structures suggests that termination of the duplex and definition of the splice site are governed by constraints of the helical geometry within the ID3 loop. This feature and flexibility of the uncomplexed ID3 loop appear to be common for both the ai5γ and O.i. introns and may help to fine-tune elements of recognition in group II introns.
通过 II 组内含子的 5' 剪接位点识别涉及到 ID3 结构域 1 茎环内的外显子结合序列（EBS）1 与外显子 1 的 3' 端互补序列（IBS1）之间的配对。为了确定 IIB 组 ai5γ 内含子剪接位点定义的分子基础，我们通过溶液 NMR 探测了 ID3 茎环单独及其与 IBS1 靶标结合时的溶液结构。ID3 茎被结构化。ID3 环的碱基被堆积，但显示出高度灵活的 EBS1 区域。EBS1 的灵活性似乎是 ai5γ 和较小的海洋芽孢杆菌（O.i.）内含子的一般特征，并且可能有助于有效搜索构象空间并防止由于偶然碱基配对而导致的剪接错误。IBS1 的结合导致形成一个结构的七个碱基对双链，尽管存在额外的 A-U 和 G•U 对的可能性，但终止于 5' 剪接位点。将这些数据与从已发表的结构中提取的 EBS1-IBS1 双链体的构象特征进行比较表明，双链体的终止和剪接位点的定义受 ID3 环内螺旋几何形状的约束。未复合的 ID3 环的这种特征和灵活性似乎对于 ai5γ 和 O.i. 内含子都是共同的，并且可能有助于微调 II 组内含子中识别元件。
