| Summary: | 碩士 === 國立臺灣大學 === 分子與細胞生物學研究所 === 105 === In prokaryotes, ribosomes have to recognize the Shine-Dalgarno sequence (SDsequence) on mRNA 5’UTR, and then the initiator tRNA can pair to the start codon to initiate translation. Many mRNAs fold into secondary structures in normal situation.During the translation initiation process, these secondary structures have to be unfolded to revel the start codon. The rpsO gene transcript 5’ untranslated region (5’UTR) of Escherichia coli regulates its own translation through folding into a doublehairpin or a pseudoknot conformation. However, the SD sequence is concealed in the double-hairpin structure. Ribosomes can only bind to pseudoknot conformation to initiate translation. The SD sequence is fully exposed only on the pseudoknot. The start codon is adjacent to the downstream secondary structure, so the ribosome has to unwind the secondary structure to complete the initiation process.
In this research, we investigate the conformational change of the rpsO 5’ UTR in the presence of the ribosomal subunit 30S and initiator tRNA by using optical tweezers. Our results showed that there was no significant difference for the rpsO 5’UTR treated with or without 30S. In the presence of the 30S and initiator tRNA, rpsO 5’ UTR downstream secondary structure could be completely unfolded. The optical tweezers’ data showed that nucleotides deletion between the two hairpins decreased the 30S binding probability to RNA molecules, and also decreased the in vitro translation efficiency. These results demonstrate that the 30S and initiator tRNA can unfold the rpsO 5’ UTR downstream secondary structure, and the length of nucleotides between the two hairpins plays an important role in binding of 30S and tRNA to rpsO 5’ UTR.
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