Journal Article

The Influence of Base Identity and Base Pairing on the Function of the α-Sarcin Loop of 23S rRNA

Michael O'Connor and Albert E. Dahlberg

in Nucleic Acids Research

Volume 24, issue 14, pages 2701-2705
Published in print July 1996 | ISSN: 0305-1048
Published online July 1996 | e-ISSN: 1362-4962 | DOI: http://dx.doi.org/10.1093/nar/24.14.2701
The Influence of Base Identity and Base Pairing on the Function of the α-Sarcin Loop of 23S rRNA

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The α-sarcin loop of large subunit rRNAs is one of the sites of interaction of elongation factors with the ribosome, and the target of the cytotoxins α-sarcin and ricin. Using a genetic selection for increased frameshifting in a reporter gene, we have isolated a C→U mutation at position 2666 in the α-sarcin loop. In the NMR-derived structure of the loop, bases equivalent to 2666 and 2654 are paired via a non-canonical base pairing interaction. Each of the three base substitutions at C2666 and A2654 was constructed by site-directed mutagenesis of a plasmid borne copy of the rrnB operon of Escherichia coli. Only the C2666→U and A2654→G mutations that resulted in the formation of canonical A-U and C-G base pairs respectively, increased the levels of stop codon readthrough and frameshifting. The effects of different base pair combinations at positions 2666 and 2654 on ribosome function were then tested by constructing and analyzing all possible base combinations at these sites. All A→ G base substitution mutations at position 2654 and C→ U substitutions at position 2666 increased the levels of translational errors. However, these effects were greatest when G2654 and U2666 had the potential to engage in standard Watson-Crick base Pairing Interactions. These Data Indicate that base Identity as well as base Pairing Interactions are Important for the Function of this Essential Component of the Large Subunit rRNA.

Journal Article.  3049 words.  Illustrated.

Subjects: Chemistry ; Biochemistry ; Bioinformatics and Computational Biology ; Genetics and Genomics ; Molecular and Cell Biology

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