The carbonic anhydrase gene families ofChlamydomonas reinhardtii

Abstract

Carbonic anhydrases (CAs) are zinc-containing metalloenzymes that catalyze the reversible interconversion of CO2 and HCO3–. Aquatic photosynthetic organisms have evolved different forms of CO2-concentrating mechanisms to aid Rubisco in capturing CO2 from the surrounding environment. One aspect of all CO2-concentrating mechanisms is the critical roles played by various specially localized extracellular and intracellular CAs. There are three evolutionarily unrelated CA families designated α-, β-, and γ-CA. In the green alga, Chlamydomonas reinhardtii Dangeard, eight CAs have now been identified, including three α-CAs and five β-CAs. In addition, C. reinhardtii has another CA-like gene, Glp1 that is similar to known γ-CAs. To characterize these different CA isoforms, some of the CA genes have been overexpressed to determine whether the proteins have CA activity and to generate antibodies for in vivo immunolocalization. The CA proteins Cah3, Cah6, and Cah8, and the γ-CA-like protein, Glp1, have been overexpressed. Cah3, Cah6, and Cah8 have CA activity, but Glp1 does not. At least two of these proteins, Cah3 and Cah6, are localized to the chloroplast. Using immunolocalization and sequence analyses, we have determined that Cah6 is located to the chloroplast stroma and confirmed that Cah3 is localized to the chloroplast thylakoid lumen. Activity assays show that Cah3 is 100 times more sensitive to sulfonamides than Cah6. We present a model on how these two chloroplast CAs might participate in the CO2-concentrating mechanism of C. reinhardtii. Key words: carbonic anhydrase, CO2-concentrating mechanism, Chlamydomonas, immunolocalization.

Crystal Structure and Functional Characterization of Photosystem II-Associated Carbonic Anhydrase CAH3 in Chlamydomonas reinhardtii

“…CrCAH3 is associated with PSII, or is at least in its close vicinity under certain conditions (Pronina and Semenenko, 1984;Karlsson et al, 1998;Villarejo et al, 2002;Mitra et al, 2005). The catalytic site of water oxidation consists of a penta-m-oxo bridged tetramanganese calcium cluster (Mn 4 CaO 5 ) positioned within the OEC of PSII (Yano et al, 2006;Umena et al, 2011).…”

Acclimation to Very Low CO2: Contribution of Limiting CO2 Inducible Proteins, LCIB and LCIA, to Inorganic Carbon Uptake in Chlamydomonas reinhardtii

“…LCIB, located around the pyrenoid, traps CO 2 , either escaping from the pyrenoid or entering from outside the cell, into the stromal HCO 3 2 pool. This stromal HCO 3 2 pool then provides substrate to the pyrenoid thylakoid tubules for CAH3-catalyzed dehydration to CO 2 for Rubisco, as described in previous CCM models (Spalding et al, 1983b;Mitra et al, 2005;Spalding, 2008;Moroney et al, 2011).…”

Proposed Carbon Dioxide Concentrating Mechanism in Chlamydomonas reinhardtii

“…Immunoblot studies using antibodies raised against CAH3 demonstrated that CAH3 is associated with the thylakoid membrane (37). More specifically, immunolocalization studies indicated that CAH3 is localized on the luminal side of the thylakoids and inside the pyrenoid tubules (47). The evidence that CAH3 plays an essential role in the CCM is persuasive.…”

“…CAH6 is a constitutively expressed ␤-CA in the chloroplast stroma (47,48). This CA might be involved in recapturing CO 2 as it effluxes from the thylakoid lumen and in helping to maintain a high concentration of inorganic carbon in the stroma.…”