Due to the essential nature of this enzyme, nature has evolved the catalytic capacity to hydrate carbon dioxide/dehydrate bicarbonate several times. There are three recognised classes of carbonic anhydrase enzymes, alpha, beta and gamma, which have no significant sequence identity, and have structurally distinct overall folds. Yet, despite their structural differences, the active sites of all three classes function with a single zinc atom that is essential for catalysis. These enzymes are of ancient origin, and appear to have evolved independently from one another, thereby providing an excellent example of convergent evolution. The three classes have differing distributions in different organisms: in mammals, all the isozymes so far discovered belong to the alpha-class; plants produce mainly the beta-class; prokaryotes encode all three classes of enzyme, with the beta- and gamma-classes predominating.
The alpha-class of enzyme has been studied the most intensively in mammals, but it also occurs in prokaryotes. This class is characterised by its high affinity for zinc, where the geometry of the conserved histidine residues favours zinc binding, and is destabilised by the binding of other metals. Certain proteins from Vaccinia and other Poxviruses appear to be related to alpha-class carbonic anhydrases, but have lost two of the zinc-binding histidines and other conserved residues; these proteins are involved in cell surface-binding, and are expressed at late times in infection.
In mammals, the different isozymes vary in their tissue and subcellular distributions and in their susceptibility to inhibitors. In addition to their involvement in pH regulation, bicarbonate reabsorption and carbon dioxide expiration, carbonic anhydrase enzymes have a multitude of different roles in mammals, being implicated in ammonia transport, bone resorption, gastric acidity, muscle contraction, gluconeogenesis, renal acidification, and normal brain development. For example, carbonic anhydrase functions as an effective attentional gate that controls signal transfer through the neural network, in addition to being involved in signal processing and memory storage.
Several isozymes have been implicated in disease states. Carbonic anhydrase II is required by the kidney for renal acidification, and its absence in an inherited syndrome leads to osteoporosis, renal tubular acidosis and cerebral calcification. Two isozymes, carbonic anhydrases IX and XII, are expressed in a variety of malignancies, and appear to be associated with poor prognosis, perhaps being indicative of an aggressive malignant phenotype. In the brain, carbonic anhydrase dysfunction impairs cognition and is associated with mental retardation, Alzheimer’s disease and aging. Treatment frequently involves the use of sulphonamides that inhibit carbonic anhydrase activity, such as for the treatment of glaucoma, epilepsy, gastro-duodenal ulcers, and possibly cancer. Activators of carbonic anhydrase might have important uses in the treatment of genetic carbonic anhydrase deficiencies and memory disorders.
The beta-class of carbonic anhydrase is found in plants, algae, bacteria and archaea, and is far more diverse in sequence than the other two classes, with only five residues (three forming the zinc ligand) being completely conserved. The beta-class enzymes can be divided into seven clades (A-G) based on sequence identity, with the plant enzymes forming two clades representing dicotyledonous and monocotyledonous plants. Enzymes within these clades can vary with respect to structure and their response to inhibitors, suggesting different functional mechanisms of action. Characterisation of these enzymes reveals sharp differences between the beta class, which forms dimers, tetramers, hexamers and octomers, and the alpha and gamma classes, which form strictly monomers and trimers.
The expression of carbonic anhydrase in prokaryotes is influenced by growth rate, and is expressed at the highest levels in slow-growing, high-density cultures. The demand for bicarbonate by bacteria is 1,000- to 10,000-fold higher than can be provided by uncatalysed hydration. Various metabolic processes require either carbon dioxide or bicarbonate. For instance, in E. coli CynT carbonic anhydrase, which is normally repressed, is required during cyanate metabolism to replenish the bicarbonate used during the bicarbonate-dependent hydrolysis of cyanate by the cyanase enzyme. In photosynthetic bacteria and plant chloroplasts, carbonic anhydrase is essential for photosynthetic carbon fixation.
The gamma class may be the most ancient form of carbonic anhydrases, having evolved long before the alpha class, to which it is more closely related than to the beta-class. The reaction mechanism of the gamma-class is similar to that of the alpha-class, even though the overall folds are dissimilar and the active site residues differ (apart from those that ligand the zinc). Gamma-class carbonic anhydrase is a zinc-bound enzyme that is produced at high level in E. coli. There is a possibility that iron and cobalt could substitute for zinc in certain archaea species, because these metals were found to exhibit greater carbon dioxide hydration rates when substituted in the enzyme over zinc-bound enzyme.