Interaction of the Bacillus stearothermophilus ribosomal protein S15 with 16 S rRNA: II. Specificity determinants of RNA-protein recognition.
Interaction of the Bacillus stearothermophilus ribosomal protein S15 with 16 S rRNA: II. Specificity determinants of RNA-protein recognition.
复制标题
嗜热脂肪芽孢杆菌核糖体蛋白 S15 与 16 S rRNA 的相互作用：II。
DOI：
10.1006/jmbi.1996.0482
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发表时间：
1996
期刊：
影响因子：
--
通讯作者：
Williamson,JR
中科院分区：
文献类型：
--
作者：
Batey,RT;Williamson,JR
S15是一种初级核糖体蛋白，与16s rRNA中心区域的三向连接特异性相互作用，其结合诱导RNA的构象变化。在相应的论文中，我们证明了S15与最小rRNA结合位点对应的61个核苷酸RNA具有高亲和力。本研究利用位点定向诱变、化学修饰干扰和硫代磷酸核糖核酸的碘印迹技术，研究了嗜热脂肪芽孢杆菌s15 - rrna相互作用中RNA的序列和结构决定因素。干扰蛋白质结合的突变和RNA修饰聚集在两个不同的区域，一个包含一个内部环，另一个包含一个三向连接。由两个A·G碱基对和一个凸起的鸟苷定义的内环对于特异性相互作用并不重要，然而，BS15与该内环上方的一个系统发育保守的G·U碱基对相互作用。在螺旋22的三向结附近，一个凸起的腺苷和两个碱基对也为BS15结合提供了重要的决定因素。化学修饰干扰还表明，在三向结上的四个高度系统发育保守的核苷酸可能形成非规范的G·G和U·A碱基对，这是BS15-rRNA相互作用所必需的。乙基化修饰干扰表明，BS15结合伴随着RNA的构象变化，包括螺旋20和22的取向彼此呈锐角。将诱变、化学修饰干扰分析和碘足迹分析提供的数据投影到三维模型上，表明BS15可能与螺旋22延伸面上的小凹槽相互作用。
S15 is a primary ribosomal protein that interacts specifically with a three-way junction in the central domain of 16 S rRNA, whose binding induces a conformational change in the RNA. In the accompanying paper, we demonstrated that S15 binds with high affinity to a 61 nucleotide RNA corresponding to the minimal rRNA binding site. Here, the sequence and structural determinants for the RNA in theBacillus stearothermophilusS15-rRNA interaction have been probed using site-directed mutagenesis, chemical modification interference, and iodine footprinting of phospho rothioate RNA. Mutations and RNA modifications that interfere with protein binding cluster in two distinct regions, one containing an internal loop and the other containing a three-way junction. The internal loop, defined by two A·G base-pairs and a bulged guanosine, is not important for the specific interaction, however, BS15 interacts with a phylogenetically conserved G·U base-pair above this internal loop. Near the three-way junction in helix 22, a bulged adenosine and two base-pairs adjacent to the junction also provide important determinants for BS15 binding. Chemical modification interference also suggests that four highly phylogenetically conserved nucleotides in the three-way junction may form non-canonical G·G and U·A base-pairs that are required for the BS15-rRNA interaction. Ethylation modification interference suggests that BS15 binding is accompanied by a conformational change in the RNA involving orientation of helices 20 and 22 at an acute angle with respect to one another. Projection of the data provided by mutagenesis, chemical modification interference analysis, and iodine footprinting onto a three-dimensional model illustrates that BS15 is likely to interact with the minor groove along an extended face of helix 22.