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Recognition of the highly conserved GTPase center of 23 S ribosomal RNA by ribosomal protein L11 and the antibiotic thiostrepton.

Abstract：

:The antibiotic thiostrepton, a thiazole-containing peptide, inhibits translation and ribosomal GTPase activity by binding directly to a limited and highly conserved region of the large subunit ribosomal RNA termed the GTPase center. We have previously used a filter binding assay to examine the binding of ribosomal protein L11 to a set of ribosomal RNA fragments encompassing the Escherichia coli GTPase center sequence. We show here that thiostrepton binding to the same RNA fragments can also be detected in a filter binding assay. Binding is relatively independent of monovalent salt concentration and temperature but requires a minimum Mg2+ concentration of about 0.5 mM. To help determine the RNA features recognized by L11 and thiostrepton, a set of over 40 RNA sequence variants was prepared which, taken together, change every nucleotide within the 1051 to 1108 recognition domain while preserving the known secondary structure of the RNA. Binding constants for L11 and thiostrepton interaction with these RNAs were measured. Only a small number of sequence variants had more than fivefold effects on L11 binding affinities, and most of these were clustered around a junction of helical segments. These same mutants had similar effects on thiostrepton binding, but more than half of the other sequence changes substantially reduced thiostrepton binding. On the basis of these data and chemical modification studies of this RNA domain in the literature, we propose that L11 makes few, if any, contacts with RNA bases, but recognizes the three-dimensional conformation of the RNA backbone. We also argue from the data that thiostrepton is probably sensitive to small changes in RNA conformation. The results are discussed in terms of a model in which conformational flexibility of the GTPase center RNA is functionally important during the ribosome elongation cycle.

journal_name

J Mol Biol

journal_title

Journal of molecular biology

authors

Ryan PC,Lu M,Draper DE

doi

10.1016/0022-2836(91)90932-v

subject

Has Abstract

pub_date

1991-10-20 00:00:00

pages

1257-68

issue

eissn

0022-2836

issn

1089-8638

pii

0022-2836(91)90932-V

journal_volume

221

pub_type

杂志文章

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Recognition of the highly conserved GTPase center of 23 S ribosomal RNA by ribosomal protein L11 and the antibiotic thiostrepton.