Ribosome pausing in amylase producing Bacillus subtilis during long fermentation

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.