Protonography, a powerful tool for analyzing the activity and the oligomeric state of the γ-carbonic anhydrase identified in the genome of Porphyromonas gingivalis

Carbonic anhydrases (CAs, EC 4.2.1.1) are metalloenzymes, mostly containing zinc within their active site, which catalyze a simple but physiologically relevant reaction in all life kingdoms, carbon dioxide hydration to bicarbonate and protons. Six CA classes (alpha, beta, gamma, delta, zeta and eta) and multiple CA isoforms evolved in organisms all over the phylogenetic tree, for facing the need to efficiently convert high amounts of CO2 to its hydration products. These enzymes are thus involved in many physiologic processes, such as photosynthesis, respiration, CO2 transport, electrolyte secretion in many tissues/organs; biosynthetic reactions (gluconeogenesis, lipogenesis, ureagenesis), etc. Recently, our group reported a new technique to assay CA activity on SDS-PAGE gels, named ` protonography' due to its similarity to zymography. By using protonography, the conversion of CO2 into protons can be visualized as a yellow band on a polyacrylamide gel. By using this technique we demonstrated the possibility to detect activity of the alpha-CA from Vibrio cholerae as well as the beta-and gamma-CAs present in Escherichia coli extracts. Furthermore, the activity of the newly discovered eta-class enzyme from Plasmodium falciparum has also been evidenced with protonography, illustrating its wide use. Here we show that protonography can be also useful to reveal the oligomeric state of the gamma-CA identified in the genome of the bacterial parasite colonizing the oral cavity, Porphyromonas gingivalis, possibly allowing for a simple and efficient diagnostic method. (C) 2015 Elsevier Ltd. All rights reserved.