A new type of carboxysomal carbonic anhydrase in sulfur chemolithoautotrophs from alkaline environments

Autotrophic bacteria are able to fix CO2 in a great diversity of habitats, even though this dissolved gas is relatively scarce at neutral pH and above. As many of these bacteria rely on CO2 fixation by ribulose 1,5-bisphospate carboxylase/oxygenase (RubisCO) for biomass generation, they must compensate for the catalytical constraints of this enzyme with CO2-concentrating mechanisms (CCMs). CCMs consist of CO2 and HCO3- transporters and carboxysomes. Carboxysomes encapsulate RubisCO and carbonic anhydrase (CA) within a protein shell and are essential for the operation of a CCM in autotrophic Bacteria that use the Calvin-Benson-Basham cycle. Members of the genus Thiomicrospira lack genes homologous to those encoding previously described CA, and prior to this work, the mechanism of function for their carboxysomes was unclear. In this paper, we provide evidence that a member of the recently discovered iota family of carbonic anhydrase enzymes (iota CA) plays a role in CO2 fixation by carboxysomes from members of Thiomicrospira and potentially other Bacteria. Carboxysome enrichments from Thiomicrospira pelophila and Thiomicrospira aerophila were found to have CA activity and contain iota CA, which is encoded in their carboxysome loci. When the gene encoding iota CA was interrupted in T. pelophila, cells could no longer grow under low-CO2 conditions, and CA activity was no longer detectable in their carboxysomes. When T. pelophila iota CA was expressed in a strain of Escherichia coli lacking native CA activity, this strain recovered an ability to grow under low CO2 conditions, and CA activity was present in crude cell extracts prepared from this strain.IMPORTANCEHere, we provide evidence that iota carbonic anhydrase (iota CA) plays a role in CO2 fixation by some organisms with CO2-concentrating mechanisms; this is the first time that iota CA has been detected in carboxysomes. While iota CA genes have been previously described in other members of bacteria, this is the first description of a physiological role for this type of carbonic anhydrase in this domain. Given its distribution in alkaliphilic autotrophic bacteria, iota CA may provide an advantage to organisms growing at high pH values and could be helpful for engineering autotrophic organisms to synthesize compounds of industrial interest under alkaline conditions. Here, we provide evidence that iota carbonic anhydrase (iota CA) plays a role in CO2 fixation by some organisms with CO2-concentrating mechanisms; this is the first time that iota CA has been detected in carboxysomes. While iota CA genes have been previously described in other members of bacteria, this is the first description of a physiological role for this type of carbonic anhydrase in this domain. Given its distribution in alkaliphilic autotrophic bacteria, iota CA may provide an advantage to organisms growing at high pH values and could be helpful for engineering autotrophic organisms to synthesize compounds of industrial interest under alkaline conditions.