Escherichia coli γ-carbonic anhydrase: characterisation and effects of simple aromatic/heterocyclic sulphonamide inhibitors

Carbonic anhydrases (CAs, EC 4.2.1.1) are ubiquitous metalloenzymes involved in biosynthetic processes, transport, supply, and balance of CO₂/HCO₃ ^- into the cell. In Bacteria, CAs avoid the depletion of the dissolved CO₂/HCO₃ ^- from the cell, providing them to the central metabolism that is compromised without the CA activity. The involvement of CAs in the survival, pathogenicity, and virulence of several Bacterial pathogenic species is recent. Here, we report the kinetic properties of the recombinant γ-CA (EcoCAγ) encoded in the genome of Escherichia coli. EcoCAγ is an excellent catalyst for the physiological CO₂ hydration reaction to bicarbonate and protons, with a k_cat of 5.7 × 10⁵ s^-1 and k_cat/K_M of 6.9 × 10⁶ M^-1 s^-1. The EcoCAγ inhibition profile with a broad series of known CA inhibitors, the substituted benzene-sulphonamides, and clinically licenced drugs was explored. Benzolamide showed a K_I lower than 100 nM. Our study reinforces the hypothesis that the synthesis of new drugs capable of interfering selectively with the Bacterial CA activity, avoiding the inhibition of the human α -CAs, is achievable and may lead to novel antibacterials.