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Related Concept Videos

Ribosome Profiling02:24

Ribosome Profiling

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 helps...

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Protein Engineering by Yeast Surface Display
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Rapid selection of high-affinity binders using ribosome display.

Birgit Dreier1, Andreas Plückthun

  • 1Department of Biochemistry, University of Zurich, Zurich, Switzerland.

Methods in Molecular Biology (Clifton, N.J.)
|November 19, 2011
PubMed
Summary

Ribosome display is an effective method for evolving high-affinity protein binders, like Designed Ankyrin Repeat Proteins (DARPins). This protocol details selection, affinity maturation, and off-rate selection for generating novel binders.

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Area of Science:

  • Molecular Biology
  • Protein Engineering
  • Biotechnology

Background:

  • Ribosome display is a powerful in vitro selection technique.
  • It enables the generation of high-affinity binders from diverse protein libraries.
  • Applications include antibody fragments and Designed Ankyrin Repeat Proteins (DARPins).

Purpose of the Study:

  • To provide a detailed protocol for ribosome display selection.
  • To explain affinity maturation and off-rate selection processes.
  • To facilitate the generation of novel high-affinity binders with specific targets.

Main Methods:

  • Utilizing ribosome display for in vitro selection of protein libraries.
  • Employing affinity maturation techniques to enhance binder affinity.
  • Implementing off-rate selection to refine binder specificity and binding kinetics.

Main Results:

  • Successful generation of high-affinity binders from diverse libraries.
  • Demonstrated applicability to antibody fragments and DARPins.
  • Achieved new target specificities in a few selection rounds.

Conclusions:

  • Ribosome display is a robust method for protein binder evolution.
  • The protocol enables efficient selection, maturation, and off-rate optimization.
  • This technique is valuable for developing binders with desired specificities and affinities.