Nakashima T, Yao M, Kawamura S, Iwasaki K, Kimura M, Tanaka I
Division of Biological Sciences, Graduate School of Science, Hokkaido University, Sapporo, Japan.
RNA. 2001 May;7(5):692-701. doi: 10.1017/s1355838201002345.
Ribosomal protein L5 is a 5S rRNA binding protein in the large subunit and plays an essential role in the promotion of a particular conformation of 5S rRNA. The crystal structure of the ribosomal protein L5 from Bacillus stearothermophilus has been determined at 1.8 A resolution. The molecule consists of a five-stranded antiparallel beta-sheet and four alpha-helices, which fold in a way that is topologically similar to the ribonucleoprotein (RNP) domain. The molecular shape and electrostatic representation suggest that the concave surface and loop regions are involved in 5S rRNA binding. To identify amino acid residues responsible for 5S rRNA binding, we made use of Ala-scanning mutagenesis of evolutionarily conserved amino acids occurring in the beta-strands and loop regions. The mutations of Asn37 at the beta1-strand and Gln63 at the loop between helix 2 and beta3-strand as well as that of Phe77 at the tip of the loop structure between the beta2- and beta3-strands caused a significant reduction in 5S rRNA binding. In addition, the mutations of Thr90 on the beta3-strand and Ile141 and Asp144 at the loop between beta4- and beta5-strands moderately reduced the 5S rRNA-binding affinity. Comparison of these results with the more recently analyzed structure of the 50S subunit from Haloarcula marismortui suggests that there are significant differences in the structure at N- and C-terminal regions and probably in the 5S rRNA binding.
核糖体蛋白L5是大亚基中的一种5S rRNA结合蛋白,在促进5S rRNA的特定构象方面发挥着重要作用。嗜热栖热放线菌核糖体蛋白L5的晶体结构已在1.8埃分辨率下测定。该分子由一个五链反平行β-折叠片和四个α-螺旋组成,其折叠方式在拓扑结构上类似于核糖核蛋白(RNP)结构域。分子形状和静电表示表明,凹面和环区参与5S rRNA结合。为了鉴定负责5S rRNA结合的氨基酸残基,我们对β-链和环区中进化保守的氨基酸进行了丙氨酸扫描诱变。β1链上的Asn37、螺旋2和β3链之间环上的Gln63以及β2和β3链之间环结构末端的Phe77的突变导致5S rRNA结合显著减少。此外,β3链上的Thr90以及β4和β5链之间环上的Ile141和Asp144的突变适度降低了5S rRNA结合亲和力。将这些结果与最近分析的嗜盐栖热菌50S亚基的结构进行比较表明,N端和C端区域的结构存在显著差异,5S rRNA结合可能也存在差异。