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Three-state equilibrium of Escherichia coli trigger factor
Holger Patzelt1, Günter Kramer, Thomas Rauch
1Zentrum für Molekulare Biologie (ZMBH), Universität Heidelberg, INF282, D-69120 Heidelberg, Germany.
Biological Chemistry
|November 28, 2002
Summary
Trigger Factor (TF) is a chaperone in E. coli that binds to ribosomes. Uncomplexed TF exists in a monomer-dimer equilibrium, while ribosome-bound TF is monomeric.
Area of Science:
- Molecular Biology
- Protein Biochemistry
- Microbiology
Background:
- Trigger Factor (TF) is the primary chaperone for nascent cytosolic proteins in E. coli.
- TF chaperone activity is linked to ribosome association, yet it exists in molar excess and partially dissociates post-lysis.
Purpose of the Study:
- To investigate the in vivo oligomeric states and ribosome binding dynamics of Trigger Factor (TF).
- To elucidate the equilibrium between monomeric and dimeric TF states and its relationship with ribosome association.
Main Methods:
- Size exclusion chromatography
- Crosslinking experiments
- Analytical ultracentrifugation
- Quantitative analysis of TF-ribosome complex formation
Main Results:
- Uncomplexed TF dimerizes with an apparent Kd of 18 microM, mediated by its N-terminal and C-terminal domains.
- TF is monomeric when associated with ribosomes.
- The dissociation constant for the TF-ribosome complex is approximately 1.2 microM.
- In vivo, most ribosomes are bound by monomeric TF, while uncomplexed TF exists predominantly as dimers.
Conclusions:
- TF exhibits a three-state equilibrium involving monomeric and dimeric forms.
- Ribosome binding stabilizes the monomeric state of TF.
- The observed equilibrium is crucial for TF's function in protein folding and cellular processes.