Enzyme-triggered gelation: targeting proteases with internal cleavage sites

Area of Science:

• Biochemistry
• Molecular Biology
• Assay Development

Background:

• Protease activity detection is crucial for biological research and diagnostics.
• Existing methods may lack selectivity or require complex procedures.
• Developing generalizable and selective protease assays remains an ongoing challenge.

Purpose of the Study:

• To describe a novel, generalizable method for detecting protease activity using gelation.
• To demonstrate the applicability of this method for selective detection of specific proteases.
• To establish a versatile platform for protease activity profiling.

Main Methods:

• Utilizing a gelator scaffold functionalized with a recognition sequence for the target protease.
• Employing a secondary protease to cleave residual residues, facilitating gelation.
• Designing and validating selective assays for matrix metalloproteinase-9 (MMP-9) and prostate-specific antigen (PSA).

Main Results:

• A generalizable gelation-based assay for protease activity detection was successfully developed.
• Selective detection of MMP-9 activity was achieved using a specific recognition sequence.
• Selective detection of PSA activity was also demonstrated using the same generalizable approach.

Conclusions:

• The described gelation-based method offers a versatile and selective approach for protease activity detection.
• This method provides a foundation for developing robust diagnostic and research tools for various proteases.
• The dual-protease strategy enhances assay specificity and signal generation.