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Related Experiment Video

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Author Spotlight: A Computational Approach to Decipher Amino Acid Preferences in Multispecific Protein-Protein Interactions

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Published on: January 26, 2024

Promiscuous Protein Binding as a Function of Protein Stability.

Ruth Cohen-Khait¹, Orly Dym², Shelly Hamer-Rogotner²

¹Department of Biomolecular Sciences, Weizmann Institute of Science, 76100 Rehovot, Israel.

Structure (London, England : 1993)

|December 7, 2017

Summary

This summary is machine-generated.

Researchers evolved TEM1 beta-lactamase to bind itself, revealing mutations that alter protein structure and reduce thermal stability. This study uncovers a mechanism for purging unwanted protein interactions.

Keywords:

TEM1 de novo protein interaction dimerization evolution mutations promiscuity protein-protein specificity thermal stability β-sheet augmentation

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Area of Science:

Protein engineering
Molecular evolution
Biochemistry

Background:

Proteins must discriminate between desired and undesired binding partners.
Understanding the evolution of protein-protein interactions is crucial for protein design and function.

Purpose of the Study:

To investigate the evolutionary mechanisms governing protein-protein interactions.
To engineer TEM1 beta-lactamase for self-binding and analyze the resulting structural and stability changes.

Main Methods:

Yeast surface display was used to select a library of TEM1 beta-lactamase variants.
X-ray crystallography was employed to determine the structural basis of the evolved interactions.
Thermal stability assays were performed on engineered mutants.

Main Results:

Three mutations enabled micromolar affinity binding between TEM1 beta-lactamase variants.
Selected mutations altered the protein fold, removing an N'-terminal helix and promoting beta-sheet-mediated interactions.
A key interface mutation (E58V) is analogous to the sickle-cell mutation.
Engineered mutations reduced the thermal stability of the protein, preventing their insertion into the wild-type.

Conclusions:

Protein fold modification, rather than direct interface complementarity, can drive self-binding.
Loss of thermal stability acts as a purging mechanism against undesirable protein interactions.
Evolutionary constraints can limit the incorporation of beneficial mutations due to stability trade-offs.