A Selective Phenyl Azo Acetate Substrate for Enhanced Carbonic Anhydrase Functional Analysis.

Area of Science:

• Biocatalysis and Environmental Science
• Enzyme Kinetics and Analysis

Background:

• Carbon dioxide (CO2) sequestration is crucial for climate change mitigation.
• Biocatalytic methods using carbonic anhydrases (CAs) are promising but limited by enzyme stability and assay tools.
• Developing robust analytical methods is essential for advancing CA-based CO2 capture.

Purpose of the Study:

• To synthesize and characterize a novel phenyl azo dye acetate substrate for improved CA activity assays.
• To address limitations in CA stability and analytical profiling for CO2 sequestration applications.

Main Methods:

• Synthesis and characterization of a novel phenyl azo dye acetate.
• Optimization of the substrate for spectrophotometric assays.
• Comparative analysis of the new substrate against conventional substrates (p-nitrophenyl acetate, indoxyl acetate) for CA activity and inhibition profiling.

Main Results:

• The novel phenyl azo dye acetate substrate demonstrated over 4.5-fold higher sensitivity compared to conventional substrates.
• The new assay exhibited substantial thermal stability and broader applicability across various hydrolases.
• Reliable kinetic and inhibition analyses of CAs were enabled by the optimized substrate.

Conclusions:

• The developed phenyl azo dye acetate provides a robust platform for evaluating carbonic anhydrase activity.
• This advancement supports the development of more efficient biocatalytic systems for carbon dioxide sequestration.
• Enhanced analytical tools are critical for optimizing enzyme performance in climate change mitigation strategies.