mScarlet: a bright monomeric red fluorescent protein for cellular imaging

Area of Science:

• Biochemistry
• Molecular Biology
• Biophysics

Background:

• Fluorescent proteins are essential tools in biological research.
• Existing red fluorescent proteins have limitations in brightness and stability.
• Development of novel fluorescent proteins is crucial for advanced imaging techniques.

Purpose of the Study:

• To engineer a novel monomeric red fluorescent protein with enhanced properties.
• To characterize the brightness, quantum yield, and fluorescence lifetime of the engineered protein.
• To evaluate the performance of the new fluorescent protein as a fusion tag and FRET acceptor.

Main Methods:

• Engineering of a monomeric red fluorescent protein (mScarlet) using a synthetic template.
• Spectroscopic screening techniques for protein characterization.
• X-ray crystallography to determine the chromophore structure.
• Assessment of performance as a fusion tag and Förster resonance energy transfer (FRET) acceptor.

Main Results:

• mScarlet exhibits record brightness and a quantum yield of 70%.
• The protein has a fluorescence lifetime of 3.9 ns.
• Crystal structure reveals a planar and rigidified chromophore.
• mScarlet outperforms existing red fluorescent proteins in fusion tag applications and FRET imaging.

Conclusions:

• mScarlet is a highly performant monomeric red fluorescent protein.
• Its superior properties make it ideal for advanced biological imaging.
• mScarlet advances the capabilities of fluorescent protein technology for research.