Identification of Intracellular Carbonic Anhydrase in Chlamydomonas reinhardtii which Is Distinct from the Periplasmic Form of the Enzyme

Abstract

A physiologically significant level of intracellular carbonic anhydrase has been idenfified in Chlamydomonas reinhardtii after lysis of the cell wall-less mutant, cw15, and two intracellular polypeptides have been identified which bind to anti-carbonic anhydrase antisera. The susceptibility of the intracellular activity to sulfonamide carbonic anhydrase inhibitors is more than three orders-of-magnitude less than that of the periplasmic enzyme, indicating that the intracellular activity was disfinct from the periplasmic form of the enzyme. When electrophoretically separated cell extracts or chloroplast stromal fractions were probed with either anti-C. reinhardtil periplasmic carbonic anhydrase antiserum or anti-spinach carbonic anhydrase antiserum, immunoreactive polypeptides of 45 kilodaltons and 110 kilodaltons were observed with both antisera. The strongly immunoreactive 37 kilodalton polypeptide due to the periplasmic carbonic anhydrase was also observed in lysed cells, but neither the 37 kilodalton nor the 110 kilodalton polypeptides were present in the chloroplast stromal fraction. These studies have identified intracellular carbonic anhydrase activity, and putative intracellular carbonic anhydrase polypeptides in Chlamydomonas reinhardtii represented by a 45 kilodalton polypeptide in the chloroplast and a 110 kilodalton form probably in the cytoplasm, which may be associated with an intracellular inorganic carbon concentrating system.The unicellular green alga, Chlamydomonas reinhardtii, has an efficient mechanism for the utilization of Ci3 which allows these cells to carry out photosynthesis at optimal rates when only very low concentrations of CO2 are available in 'Supported in part by the McKnight Foundation for N. E. T., J.V. M., and H.

Extra‐ and intra‐cellular carbonic anhydrase in relation to culture age in a high‐calcifying strain of Emiliania huxleyi Lohmann

“…The location of carbonic anhydrase external to the plasmaiemma and within the cell was similar to Chlamydomonas reinhardtii (Kimpel et al, 1983;Patel & Merrett, 1986). The intra-cellular enzyme was constitutively expressed in Chlamydomonas (Moroney et al, 1987;Husic et al, 1989) as was the chloroplast enzyine in E. huxleyi (Fig. 2).…”

“…Algae with a DIC concentrating and extra-and intra-cellular carbonic anhydrase. mechanism show a high apparent affinity for DIC (Moroney et aL, 1985) whereas at high photon flux \ \ , . ^ .,..,,..…”

“…In many microalgae external and . Measurements of bicarbonate transinternal enzymes of carbonic anhydrase (Kimpel, port in high and low calcifying cells showed a rapid Togasaki & Miyachi, 1983;Aizawa & Miyachi, equilibrium between extracellular HCOg" and the 1984;Coleman et al 1984;Moroney, Husic & internal inorganic carbon pools and in the high are an essential component of a DIC calcifying cells the rapid photosynthetic fixation of {dissolved inorganic carbon) concentrating mech-''^C derived from the internal inorganic carbon pools anism that elevates the intra-cellular COg concen-suggested a role for carbonic anhydrase (Nimer & tration around Rubisco to suppress the oxygen . In this paper we report on the inhibition of photosynthesis (Badger, Kaplan & relationship between calcification, DIG utilization Berry, 1980).…”

Amino Acid Sequence Homology between N- and C-terminal Halves of a Carbonic Anhydrase in Porphyridium purpureum, as Deduced from the Cloned cDNA

“…Isolation and sequencing of its cDNA revealed the existence of possible functional amino acid residues conserved in eukaryotic-type of CAs (13). On the other hand, although the presence and physiological importance of intracellular CA were reported in C. reinhardtii (14) and other eukaryotic microalgae (8,9), their molecular characteristics have not been elucidated. In cyanobacteria, the genomic DNA region, which complements the Synechococcus PCC7942 mutant requiring a higher concentration of inorganic carbon for growth, contains an open reading frame homologous to CAs of higher plants (6).…”

Isolation and Characterization of a Mutant of Chlamydomonas reinhardtii Deficient in the CO2 Concentrating Mechanism

“…It is presently thought to involve a C, transporter (14,20) and one or more isozymes of carbonic anhydrase (10,14,21). A carbonic anhydrase located in the periplasmic space of the alga has been identified as part of the CO2 concentrating system (6, 1 1, 31).…”