Identification and Characterization of a Periplasmic α‐Carbonic Anhydrase (CA) in the Gametophytes of Saccharina japonica (Phaeophyceae)

Periplasmic or external carbonic anhydrases (CAs) have been well accepted as playing a crucial role in the acquisition of dissolved inorganic carbon; however, no cytological evidence or molecular information on these enzymes has been reported in seaweeds to date. In this study, the full‐length cDNA sequence coding for a putative periplasmic Sjα‐CA2 was cloned from the gametophytes of Saccharina japonica , an industrial brown seaweed. It was 1,728 bp in length and included a 263‐bp 5′‐untranslated region (UTR), a 577‐bp 3′‐UTR, and an 888‐bp open reading frame encoding a protein precursor consisting of 295 amino acids. The mature protein, after removal of a predicted 28‐residue signal peptide, was composed of 267 amino acids with a relative molecular weight of 29.27 kDa. Multisequence alignment and phylogenetic analysis indicated that it was a member of the α‐CA family. Enzyme activity assays showed that the recombinant Sjα‐CA2 in Escherichia coli possessed CO 2 hydration and esterase activities, thus identifying this gene Sjα ‐ CA2 in function. Immunogold electron microscopic observations with the prepared anti‐Sjα‐CA2 polyclonal antibody illustrated that Sjα‐CA2 was located in periplasmic space of the kelp gametophyte cells. Quantitative real‐time PCR results revealed that the transcription of Sjα‐CA2 was induced by elevated HCO 3 ‐ levels, but it was little changed while the kelp gametophytes were subjected to elevated CO 2 concentrations. This study suggests that the periplasmic Sjα‐CA2 might play a role in adapting to elevated environmental levels of HCO 3 ‐ by dehydration of HCO 3 ‐ to generate CO 2 , which could be readily taken up by S . japonica gametophytes.