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Ribosomal Features Essential for tna Operon Induction: Tryptophan Binding at the Peptidyl Transferase Center

Ribosomal Features Essential for tna Operon Induction: Tryptophan Binding at the Peptidyl... Ribosomal Features Essential for tna Operon Induction: Tryptophan Binding at the Peptidyl Transferase Center ▿ Luis R. Cruz-Vera 1 , Aaron New 2 , Catherine Squires 2 , and Charles Yanofsky 1 , * 1 Department of Biological Sciences, Stanford University, Stanford, California 94305 2 Department of Molecular Biology and Microbiology, School of Medicine, Tufts University, Boston, Massachusetts 02111 ABSTRACT Features of the amino acid sequence of the TnaC nascent peptide are recognized by the translating ribosome. Recognition leads to tryptophan binding within the translating ribosome, inhibiting the termination of tnaC translation and preventing Rho-dependent transcription termination in the tna operon leader region. It was previously shown that inserting an adenine residue at position 751 or introducing the U2609C change in 23S rRNA or introducing the K90W replacement in ribosomal protein L22 prevented tryptophan induction of tna operon expression. It was also observed that an adenine at position 752 of 23S rRNA was required for induction. In the current study, the explanation for the lack of induction by these altered ribosomes was investigated. Using isolated TnaC-ribosome complexes, it was shown that although tryptophan inhibits puromycin cleavage of TnaC-tRNA Pro with wild-type ribosome complexes, it does not inhibit cleavage with the four mutant ribosome complexes examined. Similarly, tryptophan prevents sparsomycin inhibition of TnaC-tRNA Pro cleavage with wild-type ribosome complexes but not with these mutant ribosome complexes. Additionally, a nucleotide located close to the peptidyl transferase center, A2572, which was protected from methylation by tryptophan with wild-type ribosome complexes, was not protected with mutant ribosome complexes. These findings identify specific ribosomal residues located in the ribosome exit tunnel that recognize features of the TnaC peptide. This recognition creates a free tryptophan-binding site in the peptidyl transferase center, where bound tryptophan inhibits peptidyl transferase activity. http://www.deepdyve.com/assets/images/DeepDyve-Logo-lg.png Journal of Bacteriology American Society For Microbiology

Ribosomal Features Essential for tna Operon Induction: Tryptophan Binding at the Peptidyl Transferase Center

Ribosomal Features Essential for tna Operon Induction: Tryptophan Binding at the Peptidyl Transferase Center

Journal of Bacteriology , Volume 189 (8): 3140 – Apr 15, 2007

Abstract

Ribosomal Features Essential for tna Operon Induction: Tryptophan Binding at the Peptidyl Transferase Center ▿ Luis R. Cruz-Vera 1 , Aaron New 2 , Catherine Squires 2 , and Charles Yanofsky 1 , * 1 Department of Biological Sciences, Stanford University, Stanford, California 94305 2 Department of Molecular Biology and Microbiology, School of Medicine, Tufts University, Boston, Massachusetts 02111 ABSTRACT Features of the amino acid sequence of the TnaC nascent peptide are recognized by the translating ribosome. Recognition leads to tryptophan binding within the translating ribosome, inhibiting the termination of tnaC translation and preventing Rho-dependent transcription termination in the tna operon leader region. It was previously shown that inserting an adenine residue at position 751 or introducing the U2609C change in 23S rRNA or introducing the K90W replacement in ribosomal protein L22 prevented tryptophan induction of tna operon expression. It was also observed that an adenine at position 752 of 23S rRNA was required for induction. In the current study, the explanation for the lack of induction by these altered ribosomes was investigated. Using isolated TnaC-ribosome complexes, it was shown that although tryptophan inhibits puromycin cleavage of TnaC-tRNA Pro with wild-type ribosome complexes, it does not inhibit cleavage with the four mutant ribosome complexes examined. Similarly, tryptophan prevents sparsomycin inhibition of TnaC-tRNA Pro cleavage with wild-type ribosome complexes but not with these mutant ribosome complexes. Additionally, a nucleotide located close to the peptidyl transferase center, A2572, which was protected from methylation by tryptophan with wild-type ribosome complexes, was not protected with mutant ribosome complexes. These findings identify specific ribosomal residues located in the ribosome exit tunnel that recognize features of the TnaC peptide. This recognition creates a free tryptophan-binding site in the peptidyl transferase center, where bound tryptophan inhibits peptidyl transferase activity.

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Publisher
American Society For Microbiology
Copyright
Copyright © 2007 by the American society for Microbiology.
ISSN
0021-9193
eISSN
1098-5530
DOI
10.1128/JB.01869-06
pmid
17293420
Publisher site
See Article on Publisher Site

Abstract

Ribosomal Features Essential for tna Operon Induction: Tryptophan Binding at the Peptidyl Transferase Center ▿ Luis R. Cruz-Vera 1 , Aaron New 2 , Catherine Squires 2 , and Charles Yanofsky 1 , * 1 Department of Biological Sciences, Stanford University, Stanford, California 94305 2 Department of Molecular Biology and Microbiology, School of Medicine, Tufts University, Boston, Massachusetts 02111 ABSTRACT Features of the amino acid sequence of the TnaC nascent peptide are recognized by the translating ribosome. Recognition leads to tryptophan binding within the translating ribosome, inhibiting the termination of tnaC translation and preventing Rho-dependent transcription termination in the tna operon leader region. It was previously shown that inserting an adenine residue at position 751 or introducing the U2609C change in 23S rRNA or introducing the K90W replacement in ribosomal protein L22 prevented tryptophan induction of tna operon expression. It was also observed that an adenine at position 752 of 23S rRNA was required for induction. In the current study, the explanation for the lack of induction by these altered ribosomes was investigated. Using isolated TnaC-ribosome complexes, it was shown that although tryptophan inhibits puromycin cleavage of TnaC-tRNA Pro with wild-type ribosome complexes, it does not inhibit cleavage with the four mutant ribosome complexes examined. Similarly, tryptophan prevents sparsomycin inhibition of TnaC-tRNA Pro cleavage with wild-type ribosome complexes but not with these mutant ribosome complexes. Additionally, a nucleotide located close to the peptidyl transferase center, A2572, which was protected from methylation by tryptophan with wild-type ribosome complexes, was not protected with mutant ribosome complexes. These findings identify specific ribosomal residues located in the ribosome exit tunnel that recognize features of the TnaC peptide. This recognition creates a free tryptophan-binding site in the peptidyl transferase center, where bound tryptophan inhibits peptidyl transferase activity.

Journal

Journal of BacteriologyAmerican Society For Microbiology

Published: Apr 15, 2007

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