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Updated: Apr 16, 2026
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Chromatographic Purification of Highly Active Yeast Ribosomes
Published on: October 24, 2011
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Purification of 70S ribosomes
Maria C Rivera1, Bruce Maguire2, James A Lake3
1Department of Biology and Center for the Study of Biological Complexity, Virginia Commonwealth University, Richmond, Virginia 23284;
Cold Spring Harbor Protocols
|March 4, 2015
Summary
This study refines prokaryotic ribosome purification using sucrose gradient centrifugation. The method isolates intact, translationally active 70S ribosomes and subunits for cell-free systems.
Area of Science:
- Molecular Biology
- Biochemistry
- Cell Biology
Background:
- Prokaryotic ribosomes are essential for protein synthesis.
- Efficient purification of active ribosomes is crucial for biochemical studies.
- Previous methods required further refinement for isolating specific ribosomal fractions.
Purpose of the Study:
- To describe a refined purification protocol for prokaryotic ribosomal particles.
- To isolate translationally active 70S ribosomes and their subunits.
- To provide intact vacant ribosomes for cell-free translation systems.
Main Methods:
- Further purification of prokaryotic ribosomal particles via sucrose cushion centrifugation.
- Final purification step using 7%-30% linear sucrose gradient centrifugation.
- Isolation of tight couple 70S ribosomes, 30S subunits, and 50S subunits.
Main Results:
- Successfully isolated intact vacant ribosomes (tight couples).
- Separated dissociated 30S and 50S ribosomal subunits.
- Obtained a translationally active ribosome fraction.
Conclusions:
- The described method provides a highly purified fraction of active prokaryotic ribosomes.
- These purified ribosomes are suitable for use in cell-free translation systems.
- This purification strategy enhances the study of prokaryotic translation.
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