Contribution of 16S rRNA nucleotides forming the 30S subunit A and P sites to translation in Escherichia coli.
Many contacts between the ribosome and its principal substrates, tRNA and mRNA, involve universally conserved rRNA nucleotides, implying their functional importance in translation. Here, we measure the in vivo translation activity conferred by substitution of each 16S rRNA base believed to contribute to the A or P site. We find that the 30S P site is generally more tolerant of mutation than the 30S A site. In the A site, A1493C or any substitution of G530 or A1492 results in complete loss of translation activity, while A1493U and A1493G decrease translation activity by >20-fold. Among the P-site nucleotides, A1339 is most critical; any mutation of A1339 confers a >18-fold decrease in translation activity. Regarding all other P-site bases, ribosomes harboring at least one substitution retain considerable activity, >10% that of control ribosomes. Moreover, several sets of multiple substitutions within the 30S P site fail to inactivate the ribosome. The robust nature of the 30S P site indicates that its interaction with the codon-anticodon helix is less stringent than that of the 30S A site. In addition, we show that G1338A suppresses phenotypes conferred by m(2)G966A and several multiple P-site substitutions, suggesting that adenine at position 1338 can stabilize tRNA interaction in the P site.
Abdi
Nimo M
NM
Department of Microbiology, The Ohio State University, Columbus, 43210, USA.
Fredrick
Kurt
K
D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D009154:Mutation; D010442:Peptide Chain Initiation, Translational; D010641:Phenotype; D012329:RNA, Bacterial / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D012336:RNA, Ribosomal, 16S / Q000502:physiology*; D012343:RNA, Transfer / Q000235:genetics / Q000378:metabolism; D012269:Ribosomal Proteins / Q000378:metabolism*; D012270:Ribosomes / Q000378:metabolism*
D012329:RNA, Bacterial; D012333:RNA, Messenger; D012336:RNA, Ribosomal, 16S; D012269:Ribosomal Proteins; C040469:ribosomal protein S30; D012343:RNA, Transfer