High-level production in a plant system of a thermostable carbonic anhydrase and its immobilization on microcrystalline cellulose beads for CO ₂ capture

Plant-produced SazCA and its application to CO₂capture. Technologies that rely on chemical absorption or physical adsorption have been developed to capture CO₂ from industrial flue gases and sequester it at storage sites. Carbonic anhydrases (CAs), metalloenzymes, that catalyze the reversible hydration of CO₂ have recently received attention as biocatalysts in the capture of CO₂ from flue gases, but their cost presents a major obstacle for use at an industrial scale. This cost, however, can be reduced either by producing a long-lasting enzyme suitable for CO₂ capture or by lowering production costs. High-level expression, easy purification, and immobilization of CAs from Sulfurihydrogenibium azorense (SazCA) were investigated in a plant system. Fusion of the 60-amino acid-long ectodomain (M-domain) of the human receptor-type tyrosine-protein phosphatase C increased the levels of SazCA accumulation. Fusion of the cellulose-binding module (CBM3) from Clostridium thermocellum resulted in tight binding of recombinant protein to microcrystalline cellulose beads, enabling easy purification. The chimeric fusion protein, BMC-SazCA, which consisted of SazCA with the M and CBM3 domains, was expressed in tobacco (Nicotiana benthamiana), giving a recombinant protein yield in leaf extracts of 350 mg/kg fresh weight. BMC-SazCA produced in planta was active in the presence of various chemicals used in CO₂ capture. Immobilization of BMC-SazCA on the surface of microcrystalline cellulose beads extended its heat stability, allowing its reuse in multiple rounds of the CO₂ hydration reaction. These results suggest that production of SazCA in plants has great potential for CA-based CO₂ sequestration and mineralization.

Keywords: Carbonic anhydrase; Cellulose-binding module; Enzyme immobilization; Nicotiana benthamiana; Plant expression system; Reversible hydration reaction.