Modular engineering of thermoresponsive allosteric proteins

Area of Science:

• Molecular Biology
• Synthetic Biology
• Biotechnology

Background:

• Thermogenetics offers noninvasive control of protein activity but is limited to transcriptional regulation and membrane recruitment.
• Existing thermogenetic tools lack broad applicability across diverse protein functions and cellular systems.

Purpose of the Study:

• To develop a generalizable strategy for engineering thermosensitive allosteric proteins.
• To expand the applications of thermogenetics beyond current limitations.
• To create a blueprint for temperature-dependent protein control.

Main Methods:

• Engineered thermosensitive allosteric proteins by inserting optimized Avena sativa LOV2 domain variants.
• Applied the strategy to various proteins in Escherichia coli and mammalian systems.
• Incorporated a chemoreceptor domain as an alternative thermosensing module.

Main Results:

• Generated potent, thermoswitchable chimeric proteins with tight temperature control (37-41°C) in E. coli.
• Engineered CRISPR-Cas genome editors responsive to physiological temperature changes in mammalian cells.
• Demonstrated that thermosensitivity is a common feature in receptor domains.

Conclusions:

• The LOV2 domain insertion strategy provides a versatile platform for engineering thermosensitive proteins.
• This approach significantly expands the toolkit for thermogenetics, enabling precise control over diverse protein functions.
• The findings pave the way for novel applications in synthetic biology and biotechnology requiring spatiotemporal regulation.