Engineering an efficient and bright split Corynactis californica green fluorescent protein

Area of Science:

• Biochemistry
• Cellular Biology
• Molecular Biology

Background:

• Split fluorescent proteins (SFPs) are crucial tools in cellular biology for applications like protein translocation and interaction studies.
• Existing SFPs, such as split green fluorescent protein (GFP) from Aequorea victoria, have limitations, particularly in multiplexing experiments requiring non-cross-reactive and efficient protein labeling.

Purpose of the Study:

• To engineer a novel monomeric split green fluorescent protein (ccGFP) with enhanced performance for cellular labeling.
• To assess the efficiency and multiplexing capabilities of the new ccGFP compared to existing split GFPs.

Main Methods:

• Engineered a monomeric split ccGFP from a tetrameric GFP found in the coral Corynactis californica.
• Evaluated the complementation speed of split ccGFP against the original A. victoria split GFP.
• Demonstrated multiplexed labeling capabilities using split ccGFP with other split fluorescent proteins (YFP, CFP).

Main Results:

• The engineered split ccGFP exhibits up to threefold faster complementation compared to the original A. victoria split GFP.
• Split ccGFP demonstrates efficient performance in cellular labeling applications.
• The new ccGFP is compatible with existing split yellow fluorescent protein (YFP) and cyan fluorescent protein (CFP) for multiplexed labeling.

Conclusions:

• The novel monomeric split ccGFP from C. californica offers significant improvements in complementation speed and efficiency.
• Split ccGFP is a valuable tool for advanced cellular biology research, particularly for multiplexed protein labeling using CRISPR technology.
• This engineered fluorescent protein expands the toolkit for complex biological imaging and interaction studies.