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Related Experiment Video

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Building a Simple and Versatile Illumination System for Optogenetic Experiments

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Creating Red Light-Switchable Protein Dimerization Systems as Genetically Encoded Actuators with High Specificity.

Zhimin Huang¹, Zengpeng Li^1,2, Xiao Zhang¹

¹Department of Biochemistry and Institute for Protein Design, University of Washington, Seattle, Washington 98195, United States.

ACS Synthetic Biology

|November 12, 2020

Summary

This summary is machine-generated.

Researchers developed a new method (COMBINES-LID) to create red light-controlled protein dimerization systems. These novel systems, called nanoReD, use a mammalian-compatible chromophore and offer precise control for potential cell and gene therapies.

Keywords:

bacterial phytochrome combinatorial antibody library light-induced dimerization nanobody optogenetics phage display

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Spatiotemporal Control of Protein Activity through Optogenetic Allosteric Regulation

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Area of Science:

Optogenetics
Molecular Biology
Biotechnology

Background:

Red light-controlled protein dimerization is crucial for advanced therapies like cell and gene therapies.
Existing systems face challenges with exogenous chromophores and suboptimal binding proteins.
Need for efficient, generalizable systems with improved specificity and mammalian compatibility.

Purpose of the Study:

To develop a novel, generalizable method (COMBINES-LID) for creating highly specific, reversible light-induced heterodimerization systems.
To engineer red light-induced dimerization (nanoReD) systems independent of existing photoreceptor binders.
To demonstrate the utility of these systems in mammalian applications.

Main Methods:

Employed a two-step binder screening approach (phage display and yeast two-hybrid) on a nanobody library.
Selected binders that specifically engage the light-activated form of a photoswitchable protein.
Engineered nanoReD systems using a truncated bacterial phytochrome and the endogenous chromophore biliverdin.

Main Results:

Successfully engineered nanoReD systems with low dark activity and high induction specificity.
Demonstrated reversible control of protein translocation in mammalian cells.
Showcased activation of gene expression in vivo (in mice).

Conclusions:

The COMBINES-LID method provides an efficient and generalizable platform for creating novel light-controlled dimerization systems.
nanoReD systems offer precise optical control over biological processes, applicable to cell and gene therapies.
This work expands the toolkit for genetically encoded actuators for optical manipulation of biological functions.