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Transcription Attenuation in Prokaryotes02:42

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Transcriptional attenuation occurs when RNA transcription is prematurely terminated due to the formation of a terminator mRNA hairpin structure.  Bacteria use these hairpins to regulate the transcription process and control the synthesis of several amino acids including histidine, lysine, threonine, and phenylalanine. Transcription attenuation takes place in the non-coding regions of mRNA.
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Base complementarity between the three base pairs of mRNA codon and the tRNA anticodon is not a failsafe mechanism. Inaccuracies can range from a single mismatch to no correct base pairing at all. The free energy difference between the correct and nearly correct base pairs can be as small as 3 kcal/ mol. With complementarity being the only proofreading step, the estimated error frequency would be one wrong amino acid in every 100 amino acids incorporated. However, error frequencies observed in...
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Ribosome profiling or ribo-sequencing is a deep sequencing technique that produces a snapshot of active translation in a cell. It selectively sequences the mRNAs protected by ribosomes to get an insight into a cell’s translation landscape at any given point in time.
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The large ribosomal subunit has several important structures essential to translation. These include the peptidyl transferase center (PTC) - which is the site where the peptide bond is formed - and a large, internal, water-filled tube through which the nascent polypeptide moves. This latter structure is called the Peptide Exit Tunnel, and it begins at the PTC and spans the body of the large ribosomal subunit. During translation, as the nascent polypeptide chain is synthesized, it passes through...
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Updated: May 30, 2025

Isolation of Ribosome Bound Nascent Polypeptides in vitro to Identify Translational Pause Sites Along mRNA
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Ribosome pausing in amylase producing Bacillus subtilis during long fermentation.

Yaozu Han1, Biwen Wang1, Alberto Agnolin1

  • 1Swammerdam Institute for Life Sciences, University of Amsterdam, Science Park 904, Amsterdam, 1098 XH, The Netherlands.

Microbial Cell Factories
|January 26, 2025
PubMed
Summary
This summary is machine-generated.

Ribosome pausing was observed in Bacillus subtilis during α-amylase production, suggesting mRNA structures may cause these translation stalls. This finding aids future codon optimization for improved enzyme yields.

Keywords:
Bacillus subtilisAmylaseProtein secretionRibosome pausingRibosome profiling

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Area of Science:

  • Molecular Biology
  • Biotechnology
  • Microbial Fermentation

Background:

  • Ribosome pausing significantly impacts protein synthesis efficiency.
  • Optimizing translation is crucial for industrial applications.
  • Investigated ribosome pausing during α-amylase (AmyM) production in Bacillus subtilis.

Purpose of the Study:

  • To determine if ribosome pausing occurs during AmyM production in Bacillus subtilis.
  • To identify genome-wide ribosome pausing sites under repeated batch fermentation.

Main Methods:

  • Utilized ribosome profiling with mupirocin for codon resolution.
  • Analyzed B. subtilis ribosome pausing at 16h and 64h growth.
  • Identified pausing sites in highly expressed amyM and toxin genes.

Main Results:

  • Detected strong, persistent ribosome pausing sites in the amyM gene.
  • Pausing sites were not linked to proline, rare codons, or protein structure.
  • Identified significant pausing in several toxin genes, potentially regulating translation.

Conclusions:

  • Ribosome pausing events are associated with amyM gene expression in B. subtilis.
  • Secondary mRNA structures are hypothesized to cause pausing, not codon bias or protein structure.
  • Provides data for future codon optimization to enhance amylase production.