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Related Concept Videos

Ribosome Profiling02:24

Ribosome Profiling

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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.
Applications of ribosome profiling
Ribosome profiling has many applications, including in vivo monitoring of translation inside a particular organ or tissue type and quantifying new protein synthesis levels.
The technique...
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Ribosomes01:27

Ribosomes

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Ribosomes translate genetic information encoded by messenger RNA (mRNA) into proteins. Both prokaryotic and eukaryotic cells have ribosomes. Cells that synthesize large quantities of protein—such as secretory cells in the human pancreas—can contain millions of ribosomes.
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Ribosomes are composed of ribosomal RNA (rRNA) and proteins. In eukaryotes, rRNA is transcribed from genes in the nucleolus—a part of the nucleus that specializes in ribosome...
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Improving Translational Accuracy02:07

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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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Ribosomal RNA Synthesis02:53

Ribosomal RNA Synthesis

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Ribosome synthesis is a highly complex and coordinated process involving more than 200 assembly factors. The synthesis and processing of ribosomal components occurs not only in the nucleolus but also in the nucleoplasm and the cytoplasm of eukaryotic cells.
Ribosome biogenesis begins with the synthesis of 5S and 45S pre-rRNAs by distinct RNA polymerases. The primary transcripts are extensively processed and modified before they are bound and folded by ribosomal proteins and assembly factors,...
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Leaky Scanning02:28

Leaky Scanning

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During most eukaryotic translation processes, the small 40S ribosome subunit scans an mRNA from its 5' end until it encounters the first start AUG codon. The large 60S ribosomal subunit then joins the smaller one to initiate protein synthesis. The location of the translation initiation is largely determined by the nucleotides near the start codon as there may be multiple translation initiation sites present on the mRNA.  Marilyn Kozak discovered that the sequence RCCAUGG (where R...
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Post-translational Translocation of Proteins to the RER01:27

Post-translational Translocation of Proteins to the RER

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A sizable fraction of proteins destined for ER are first synthesized in the cell cytosol and then transported across the ER membrane–a process called post-translational translocation. Similar to cotranslationally translocated proteins, these proteins also use the Sec translocon complex to enter the ER lumen.
Targeting proteins to the ER
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Updated: Jun 27, 2025

Engineering 'Golden' Fluorescence by Selective Pressure Incorporation of Non-canonical Amino Acids and Protein Analysis by Mass Spectrometry and Fluorescence
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Engineering 'Golden' Fluorescence by Selective Pressure Incorporation of Non-canonical Amino Acids and Protein Analysis by Mass Spectrometry and Fluorescence

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Ribosome Pool Engineering Increases Protein Biosynthesis Yields.

Camila Kofman1, Jessica A Willi1, Ashty S Karim1

  • 1Department of Chemical and Biological Engineering, Northwestern University, Evanston, Illinois 60208, United States.

ACS Central Science
|April 29, 2024
PubMed
Summary
This summary is machine-generated.

Bacterial ribosomes have sequence variations impacting protein production. Engineering these ribosomes, using specific ribosomal RNA (rRNA) operons, can significantly enhance protein yields for synthetic biology and biomanufacturing.

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Engineering 'Golden' Fluorescence by Selective Pressure Incorporation of Non-canonical Amino Acids and Protein Analysis by Mass Spectrometry and Fluorescence
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Global Identification of Co-Translational Interaction Networks by Selective Ribosome Profiling
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Global Identification of Co-Translational Interaction Networks by Selective Ribosome Profiling

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Area of Science:

  • Molecular Biology
  • Synthetic Biology
  • Bacterial Genetics

Background:

  • Bacterial ribosomes, crucial for protein synthesis, possess diverse ribosomal RNA (rRNA) operon sequences.
  • Functional differences arising from these native sequence variations are not well understood.
  • Understanding these differences is key to optimizing ribosomes for synthetic biology applications.

Purpose of the Study:

  • To investigate the functional impact of sequence polymorphisms in *Escherichia coli* rRNA operons on ribosome performance.
  • To assess the protein production capabilities of ribosomes derived from different rRNA operon combinations.
  • To explore the potential of engineered ribosome pools for improving protein biomanufacturing yields.

Main Methods:

  • Utilized an *in vitro* ribosome synthesis and translation platform to analyze ribosomes from unique combinations of 16S and 23S rRNAs from seven *E. coli* rRNA operons.
  • Generated bacterial strains expressing single rRNA operons for *in vivo* analysis.
  • Assessed protein production yields using a panel of medically and industrially relevant proteins.

Main Results:

  • Sequence variations in native *E. coli* rRNA operons significantly alter ribosome function and protein production efficiency.
  • Some purified *in vivo* expressed homogeneous ribosome pools demonstrated superior performance compared to the wild-type heterogeneous pool.
  • A cell lysate expressing only operon A ribosomes showed substantial yield increases for target proteins.

Conclusions:

  • Native rRNA operon sequence polymorphisms directly influence ribosome functionality and protein synthesis.
  • Engineering ribosome pools by selecting specific rRNA operons can enhance protein yields in biomanufacturing.
  • Ribosome pool engineering offers a novel strategy for optimizing protein production and understanding ribosome evolution.