Isolation, purification, characterization and mass spectroscopic analysis of carbonic anhydrase from Serratia sp. for sequestration of carbon dioxide and formation of calcite

• Microbial community isolated from marble rock for evaluation of sequestration of carbon dioxide.
• Serratia sp., an Enterobacteriaceae, isolated from enriched microbial community had CO2 sequestration potency reported first time.
• Sequestration potency was evaluated by production of carbonic which is purified and characterized indicating similarity from mining drainage.
• Serratia sp. formed calcium carbonate indicating biosequestration of carbon dioxide through mineralization processes.

Bacterial community isolated from marble rock was enriched in the chemostat by continuous culture in minimal salt medium with different concentrations of sodium bicarbonate. One of the isolate survived at 100 mM sodium bicarbonate exhibited capability to sequester carbon dioxide (CO2) was identified as Serratia sp. by 16S rRNA sequencing. CO2 sequestering capacity was detected by carbonic anhydrase enzyme activity and formation of calcite. The carbonic anhydrase enzyme from Serratia sp. was detected after isolation, partial purification by ammonium sulfate precipitation, gel filtration chromatography, affinity chromatography, sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and MALDI-TOF/MS. The purified enzyme molecular weight was 29 kDa on SDS-PAGE and gel filtration chromatography. The optimum pH and temperature of the enzyme were found to be 7.5 and 40 °C respectively. The Km of CA was 2.25 mM and the Vmax 940 μmol/min/mg. The Kcat and Kcat/Km value of purified carbonic anhydrase was 2.8 × 105 s−1 and 1.2 × 108 M−1 s−1 respectively. MALDI-TOF mass spectrometry data showing highest matched sequence in database was similar to carbonic anhydrase. Scanning electron microscopy (SEM) analysis revealed the formation of calcium carbonate. Major calcite phase diffraction peaks occurred at 2θ = 29.4°, 36.1°, 39.5° and 43.5° determined by X-ray diffraction (XRD) analysis. Energy-dispersive X-ray (EDX) analysis showed the presence of CaO (72%) and carbon (18%) indicated role of carbon concentrating mechanisms of Serratia sp., a member of Enterobacteriaceae isolated from marble mines.