Carbonic anhydrase is extremely convergent and may have evolved as many as six times. The most familiar variants are α, β and γ carbonic anhydrases.

Carbonic anhydrase is one of the key enzymes in life. Its principal function is to engineer the hydration of carbon dioxide (or its reversible reaction to bicarbonate). It plays, therefore, a central role in facilitating the diffusion of carbon dioxide in photosynthesis, but it is also essential in areas such as respiration (with the transport of carbon dioxide in the blood), ion balance and an important role in kidney function, acid-base balance, biomineralization, and various aspects of bacterial metabolism (e.g. cyanates). It is not only a remarkably fast enzyme, but so central is it to life that one’s automatic assumption is that it would be one of the first enzymes to evolve and so employed in diverse circumstances as and when required. Nothing could be further from the truth: carbonic anhydrase is extremely convergent and may have evolved as many as six times.

Forms of carbonic anhydrase

The most familiar variants of this enzyme are α, β and γ carbonic anhydrases (CA). α-CA is the only type to be found in vertebrates, and has been extensively studied in mammals where it occurs in numerous isoforms. Malfunction of α-CA can have serious medical consequences. Although there is a rough taxonomic division of carbonic anhydrases, with β-CA for example being characteristic of plants (where it too is found as many isoforms) and γ-CA of chemolithoautotrophic bacteria, in reality the distribution of each type is generally widespread. Moreover, in a number of organisms two different types of carbonic anhydrase occur. In some coccolithophorids, a group of marine algae with characteristic calcareous plates, γ-CA occurs with δ-CA, yet another convergent form of CA. By and large it is not clear why different carbonic anhydrases co-exist, but there is little doubt they have specific functions.

α-carbonic anhydrase and ionic balance in Dunaliella

The role of mammalian α-CA in the kidney reveals another sort of convergence, striking in its own way, with a remarkable alga (Dunaliella) which is highly tolerant of salty water; indeed it can live in saturated brine. Incidentally it avoids osmotic catastrophe by adding glycerol to its cell, but in terms of its carbonic anhydrase it possesses two forms. One form performs the usual function of ensuring a supply of carbon dioxide for photosynthesis. The second form, however, has several specific adaptations, including a loop on the protein to bind sodium (Na). This second carbonic anhydrase is strikingly similar to the one found in the kidney where it performs an important function in ion balance, and also turns out (unsurprisingly) to be remarkably halotolerant. Although both carbonic anhydrases in the alga and kidney belong to the α-CA class, they have arrived necessarily at the same solution to deal with sodium.

Cadmium-based carbonic anhydrase in diatoms

Speaking of convergence in carbonic anhydrase Liljar and Lauberg (in 2000) aptly wrote of it as “A wheel invented three times”, referring to the canonical α-CA, β-CA and γ-CA. As already mentioned δ-CA is also identified as yet another distinct form, apparently having a separate origin and most typical of such algae as diatoms. What of the other two? A fifth variety is particularly interesting because in contrast to all other carbonic anhydrases it uses an atom of cadmium rather than zinc at the active site (cobalt may also serve as a substitute). It too occurs in diatoms, and may be the clue to their extraordinary success. Indeed diatoms probably sequester about a quarter of all inorganic carbon each year; remember that amongst that inorganic carbon is anthropogenic carbon dioxide! The success (and importance) of diatoms may have resulted because of the intense competition for nutrients in the ocean, especially zinc. By substituting cadmium, which in other circumstances can be toxic, this may have given this group of algae the decisive competitive advantage.

ε-carbonic anhydrase: β-like active site

The last carbonic anhydrase, ε-CA, is especially interesting. It is typical of certain bacteria, such as Halothiobacillus, and when first discovered was interpreted as a new type. More recently it has been regarded as a special and distinct variety of β-CA. Why is this? This protein has no sequence similarity and possesses a distinctive domain that is not found in β-CA. However, the domain around the active site is similar in both β- and ε-CA, and the active site is identical. However, unlike β-CA the characteristic duplication of the active site is not seen. ε-CA, therefore, is tantalizing: it clearly has similarities to β-CA, but to my mind this could as easily be convergent.

Active site constraints and molecular convergence

And why do we speak of convergence? The answer is, of course, that the active site of all carbonic anhydrase enzymes is effectively identical: using zinc (or sometimes the chemically similar cadmium) in association with three key amino acid residues: in α-CA, γ-CA and δ-CA they are three histidines, and in β-CA and ε-CA there is one histidine and two cysteines. In addition, a fourth adjacent site is typically occupied by either water or aspartate. Interestingly in the case of α-CA and β-CA the active site arrangement is a mirror-image of one another.

Carbonic anhydrase exemplifies molecular convergence. An identical function (although interestingly carbonic anhydrase is also known to be co-opted for non-catalytic functions) is achieved by the same active site, but the molecular architecture of the rest of the protein clearly shows that they have evolved independently and cannot possibly have come from a common ancestor. In at least the case of carbonic anhydrase it is surely a universal molecule: wherever carbon based life forms need to hydrate carbon dioxide, there we will find this enzyme.