| Abstract [eng] |
Carbonic anhydrases (CAs, EC 4.2.1.1) are metalloenzymes that catalyse the reversible hydration of carbon dioxide and, in some cases, the hydrolysis of esters, phosphates, and sulfates. Their high catalytic efficiency makes them promising biocatalysts for CO2 capture, conversion, and utilisation in sustainable biotechnological applications. Despite extensive studies on CA structure and medicinal applications, a comprehensive comparison of functional assays for CA activity is lacking. This concise review systematically presents and critically evaluates methods for CA activity analysis, including hydration- and hydrolysis-based approaches such as manometry, gravimetry, protonography, potentiometry, spectrophotometry, isotope exchange, and gas-based detection techniques. Each method is evaluated based on its principles, sensitivity, equipment requirements, applicability across various CA classes, and suitability for high-throughput screening. Special attention is given to methodological challenges affecting accuracy, reproducibility, and the study of novel or engineered CA variants, including those from extremophilic organisms. By consolidating existing techniques and highlighting their advantages and limitations, the review provides a practical framework for selecting and optimising CA assays. Moreover, it supports the discovery, functional characterisation, and industrial application of CAs as versatile biocatalysts, promoting CO2 valorisation and advancing sustainable processes in biocatalysis and agricultural biotechnology. |
