Deep-sea bacterial alpha carbonic anhydrase as an efficient biocatalyst for sustainable CO₂ mineralization

The ocean sequestrates carbon dioxide through bicarbonate formation and acts as a natural carbon sink. Microbial carbonic anhydrases are ubiquitous metalloenzymes that catalyze the reversible hydration of CO₂. α-carbonic anhydrases (α-CAs) are of particular interest due to their high catalytic activity in reversible hydration of CO₂, yet recombinant deep-sea CAs remain poorly characterized. The present study identified an active alpha class carbonic anhydrase producing Cytobacillus oceanisediminis SBA2, from the Arabian Sea seamount sediment. CoACA was cloned and expressed in soluble fraction of E. coli Rosetta pLysS with an IPTG induction of 0.3 mM at 20 °C for 16 h. The apparent molecular weight of purified CoACA was observed as 31.0 kDa confirmed from SDS-PAGE. CoACA existed as dimer with a specific activity of 2202 ± 75 WAU/mg, determined by CO₂ hydration assay. The enzyme retained 42% of residual activity after 1 hr of 50 bar pressure treatment and 140% activity at 200 mM salt concentration. The potential of CO₂ mineralization by purified CoACA was confirmed by calcium carbonate precipitation. Stability of the purified recombinant CoACA under high pressure revealed its piezotolerant nature compared to the commercial bovine carbonic anhydrase. The pressure tolerant CoACA could be further explored for CO₂ capture and assisted technologies.