Small Molecule-Induced Domain Swapping as a Mechanism for Controlling Protein Function and Assembly

Area of Science:

• Biochemistry
• Molecular Biology
• Protein Engineering

Background:

• Domain swapping is a protein dimerization mechanism.
• Protein function regulation is crucial in biological systems.
• Rational design of protein switches is an emerging field.

Purpose of the Study:

• Introduce induced domain swapping (INDOS) as a novel mechanism for protein function regulation.
• Demonstrate the rational design of a small molecule-inducible protein switch.
• Engineer proteins that regain function through domain swapping.

Main Methods:

• Fusion of a recognition protein (FK506 binding protein) with functionally-inactivated target proteins (staphylococcal nuclease, ribose binding protein).
• Induction of domain swapping via small molecule (FK506) binding to the recognition domain.
• Analysis of protein refolding, dimerization, and functional recovery.

Main Results:

• FK506 binding induced target protein unfolding and refolding via domain swapping.
• Inactivating mutations were 'swapped out' in the dimer.
• Nuclease activity increased 100-fold and ribose binding activity increased 15-fold, nearing wild-type levels.

Conclusions:

• INDOS provides a controllable mechanism to regulate protein function using small molecules.
• This study presents the first rational design of a small molecule-triggered protein domain swapping system.
• INDOS enables the conversion of arbitrary proteins into functional, switchable biological components.