Quantitative measurement of cell-surface displayed proteins based on split-GFP assembly

Area of Science:

• Biotechnology
• Molecular Biology
• Microbiology

Background:

• Microbial cell surface display immobilizes proteins for enhanced functionality.
• Assessing and quantifying displayed proteins on microbial cells is challenging.
• Split-green fluorescent protein (GFP) offers a potential solution through complementation-based fluorescence.

Purpose of the Study:

• To develop and validate split-GFP based methods for confirming protein display on microbial cell surfaces.
• To quantify the number of proteins displayed on individual microbial cells.
• To simplify and improve the analysis of microbial cell surface display technology.

Main Methods:

• Utilized two split-green fluorescent protein (split-GFP) assembly systems: GFP1-10/GFP11 and GFP1-9/GFP10-11.
• Evaluated display efficiency of SUMO and ubiquitin using various anchor proteins.
• Measured functional protein display (laccase) using the split-GFP systems.

Main Results:

• Successfully demonstrated the feasibility of both split-GFP assembly systems for confirming and quantifying protein display.
• Showcased the evaluation of display efficiency for different proteins and anchor proteins.
• Validated the application of split-GFP systems for measuring functional protein expression.

Conclusions:

• Developed two simple, cost-effective split-GFP methods for qualitative and quantitative analysis of cell surface display.
• These methods facilitate comprehensive investigations into microbial cell surface display technology.
• The proposed systems simplify fluorescence analysis for surface-displaying cells.