Nucleic Acids Research, Vol 24, Issue 14 2701-2705, Copyright © 1996 by Oxford University Press
M O'Connor and AE Dahlberg
The alpha-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 alpha-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 alpha-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.
ARTICLES
The influence of base identity and base pairing on the function of the alpha-sarcin loop of 23S rRNA
Department of Molecular and Cell Biology and Biochemistry, J. W. Wilson Laboratory, Brown University, Providence, RI 02912, USA.
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