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Rounding up: Engineering 12-membered rings from the cyclic 11-mer TRAP
Jonathan G Heddle1, Takeshi Yokoyama, Ichiro Yamashita
1Protein Design Laboratory, Yokohama City University, Tsurumi, Suehiro 1-7-29, Yokohama 230-0045, Japan. jgh4@tsurumi.yokohama-cu.ac.jp
Structure (London, England : 1993)
|May 16, 2006
Summary
Engineered 12-subunit TRAP protein rings bind ligands, revealing that protein-protein interactions favor strained conformations due to entropic factors, not just stability.
Area of Science:
- Protein engineering
- Structural biology
- Biophysics
Background:
- The trp RNA binding attenuation protein (TRAP) naturally forms a stable 11-subunit ring structure.
- Understanding the factors governing protein self-assembly and quaternary structure is crucial in molecular biology.
Purpose of the Study:
- To engineer novel, larger ring structures of TRAP protein.
- To investigate the conformational preferences and stability of engineered protein rings.
- To elucidate the role of entropic factors in protein-protein interactions.
Main Methods:
- Genetic engineering to create tandem DNA sequences encoding TRAP monomers.
- Protein expression and purification.
- X-ray crystallography to determine the structure of engineered 12-mer rings.
- Structural comparison between 11-mer and 12-mer TRAP rings.
Main Results:
- Successfully engineered and characterized 12-subunit TRAP rings capable of binding 12 ligand molecules.
- Crystal structures revealed conserved hydrogen bonding and buried surface areas between subunits in both 11-mer and 12-mer rings.
- Engineered 12-mer rings exhibited significant steric strain from peptide linkers and reduced thermostability compared to the native 11-mer.
Conclusions:
- Proteins can adopt strained conformations in engineered structures, indicating the significant influence of entropic contributions on protein assembly.
- The preference for forming 12-mer rings over unstrained 11-mer structures highlights the critical role of entropy in driving protein-protein interactions.
- This study provides insights into the principles of protein design and the thermodynamics of protein complex formation.
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