Carbonic Anhydrase

Carbonic anhydrase (CA) catalyzes the reversible hydration of dissolved CO2 via a two-step mechanism that involves an attack of zinc-bound OH- on a CO2 molecule loosely bound in a hydrophobic pocket. The resulting zinc-coordinated HCO3- ion is displaced from the metal ion by H2O [25]. The overall reaction is as follows:

CO2 + H2O о H+ + HCO3- (3)

Although this reaction occurs without CA, the uncatalyzed hydration and dehy­dration reactions are slow. Therefore, CA is important when the availability of CO2 or HCO3- becomes limiting to a metabolic reaction [43]. Because the reaction shown in (3) involves protons, the equilibrium ratio of the two carbon forms is a function of pH. At physiological ionic strengths, CO2 predominates at a pH of less than 6.4, and HCO3- is the dominant form in the pH range of 6.4 and 10.3.

Three main evolutionarily distinct families of CAs were initially identified (a, b, and g-CAs), and recently two more CA classes, the 5 and Z classes, have been found in marine diatoms [44]. All CA families require zinc at the active site to activate the water molecule, although the g-CAs have been shown to use iron at the active site under anaerobic conditions. However, there is no significant sequence homology between families, and they appear to be examples of convergent evolution of cata­lytic function.

2.2.1 Types of CA Present in R. eutropha

a-CA: The best-studied group of the CAs is the a-class, first described and mostly studied in mammals, but also found in other organisms [24, 26]. Most a-CAs are monomeric enzymes, with an active site zinc ion coordinated by three histidine resi­dues. Members of this family may be involved in maintaining pH balance, in facili­tating transport of carbon dioxide or bicarbonate, or in sensing carbon dioxide levels in the environment [25, 45].

b-CA: The b-class has been identified in plants, Bacteria, red and green algae, fungi, andArchaea [46,47]. Formanyorganisms, including R. eutropha, b-CAis essential for growth at atmospheric concentrations of CO2 [47, 48]. The fundamental struc­ture of b-CA, a dimer, is the only one known to exhibit allosteric regulation by bicarbonate ion [46] .

y-CA: The g-CAs, which are predominantly found in Archaea, have strikingly dif­ferent sequence features than the a — and b-CAs, The g-CAs. are trimeric enzymes that contain a zinc in the active site. In an anaerobic environment, the zinc is replaced by iron as the physiologically relevant active site metal [45, 49]. R. eutropha is capable of growth under anaerobic conditions, and may utilize a g-CA (see below) under these conditions.

Although the primary structure and number of subunits of the various classes of CAs are strikingly different, the metal — (in most cases zinc) coordinating site is remarkably similar at the structural level [45, 46].

Four putative CA genes were identified in the genome sequence of R. eutropha strain H16. Two are located on chromosome 1, and two on chromosome 2. H16_ A1192 encodes a g-like-CA/acetyltransferase, can (locus tag H16_A0169) and can2 (locus tag H16_B2270) encode b-CA enzymes, and caa (locus tag H16_B2403) encodes a periplasmic a-CA. The presence of genes for multiple carbonic anhy — drases in R. eutropha suggests that these enzymes play a major role in its physiology and that the function of the different types is complementary [24] . However, the exact roles of all four CA enzymes are still largely unknown. The only R. eutropha CA gene studied to date is can [48], which was identified as being essential for growth under atmospheric concentrations of CO2. Either the presence of the other three CAs was not sufficient to support growth under ambient CO2 concentrations, or these other CA genes were not expressed [48]. The metabolic processes in which the activity of the other three CAs plays a role remain to be identified.

Since CO2 and HCO3- are both involved in a wide range of cellular processes, CAs can have different physiological roles. One role is to increase the supply of CO2 or HCO3- for metabolic reactions. For example, during the carboxylation reaction of RuBisCO, which uses only CO2, competition between CO2 and O2 for the active site is attenuated when the concentration of CO2 is higher. It is mainly for this purpose that CA activity, and enhancement thereof, may be crucial for autotrophic IBT pro­duction in R. eutropha. Other roles CA may play include delivery of carbon to the correct location within the cell and retention of CO2. Both of these roles are impor­tant, as CO2 can readily pass through biological membranes and quickly leak out of the cell. The role of CAs in transportation and retention of CO2 is shown through the carboxysomes, or CO2 concentration mechanism (CCM), which are important in cyanobacteria and algal carbon fixation [50, 51] . The action of CAs may also be important in pH homeostasis [43]. The roles of CA enzymes in IBT production are likely to prove important, both for concentrating CO2 to drive the RuBisCO reaction towards carbon fixation and for increasing CO2 tolerance.

Добавить комментарий

Ваш e-mail не будет опубликован. Обязательные поля помечены *