Chubiz Lon M, Rao Christopher V
Department of Chemical and Biomolecular Engineering, University of Illinois at Urbana-Champaign, 600 S. Mathews Ave, Urbana, IL 61801, USA.
Nucleic Acids Res. 2008 Jul;36(12):4038-46. doi: 10.1093/nar/gkn354. Epub 2008 Jun 3.
Orthogonal ribosomes (o-ribosomes), also known as specialized ribosomes, are able to selectively translate mRNA not recognized by host ribosomes. As a result, they are powerful tools for investigating translational regulation and probing ribosome structure. To date, efforts directed towards engineering o-ribosomes have involved random mutagenesis-based approaches. As an alternative, we present here a computational method for rationally designing o-ribosomes in bacteria. Working under the assumption that base-pair interactions between the 16S rRNA and mRNA serve as the primary mode for ribosome binding and translational initiation, the algorithm enumerates all possible extended recognition sequences for 16S rRNA and then chooses those candidates that: (i) have a similar binding strength to their target mRNA as the canonical, wild-type ribosome/mRNA pair; (ii) do not bind mRNA with the wild-type, canonical Shine-Dalgarno (SD) sequence and (iii) minimally interact with host mRNA irrespective of whether a recognizable SD sequence is present. In order to test the algorithm, we experimentally characterized a number of computationally designed o-ribosomes in Escherichia coli.
正交核糖体（o-核糖体），也被称为特殊核糖体，能够选择性地翻译宿主核糖体无法识别的信使核糖核酸（mRNA）。因此，它们是研究翻译调控和探究核糖体结构的有力工具。迄今为止，构建o-核糖体的努力涉及基于随机诱变的方法。作为一种替代方法，我们在此提出一种在细菌中合理设计o-核糖体的计算方法。该算法基于16S核糖体RNA（rRNA）与mRNA之间的碱基对相互作用是核糖体结合和翻译起始的主要模式这一假设，列举出16S rRNA的所有可能的扩展识别序列，然后选择那些满足以下条件的候选序列：（i）与目标mRNA的结合强度与经典野生型核糖体/mRNA对相似；（ii）不与具有野生型经典夏因-达尔加诺（SD）序列的mRNA结合；（iii）无论是否存在可识别的SD序列，与宿主mRNA的相互作用最小。为了测试该算法，我们在大肠杆菌中对一些通过计算设计的o-核糖体进行了实验表征。
