Enzyme mediated transformation of CO₂ into calcium carbonate using purified microbial carbonic anhydrase

In this study, a bacterial carbonic anhydrase (CA) was purified from Corynebacterium flavescens for the CO₂ conversion into CaCO₃. The synthesized CaCO₃ can be utilized in the papermaking industry as filler material, construction material and in steel industry. Herein, the CA was purified by using a Sephadex G-100 column chromatography having 29.00 kDa molecular mass in SDS-PAGE analysis. The purified CA showed an optimal temperature of 35 °C and pH 7.5. In addition, a kinetic study of CA using p-NPA as substrate showed V_max (166.66 μmoL/mL/min), K_m (5.12 mM), and K_cat (80.56 sec⁻¹) using Lineweaver Burk plot. The major inhibitors of CA activity were Na²⁺, K⁺, Mn²⁺, and Al³⁺, whereas Zn²⁺ and Fe²⁺ slightly enhanced it. The purified CA showed a good efficacy to convert the CO₂ into CaCO₃ with a total conversion rate of 65.05 mg CaCO₃/mg of protein. In silico analysis suggested that the purified CA has conserved Zn²⁺ coordinating residues such as His 111, His 113, and His 130 in the active site center. Further analysis of the CO₂ binding site showed conserved residues such as Val 132, Val 142, Leu 196, Thr 197, and Val 205. However, a substitution has been observed where Trp 208 of its closest structural homolog T. ammonificans CA is replaced with Arg 207 of C. flavescens. The presence of a hydrophilic mutation in the CO₂ binding hydrophobic region is a further subject of investigation.