Carbonic anhydrase activators: The first activation study of the human secretory isoform VI with amino acids and amines Original Research Article

The secretory isozyme of human carbonic anhydrase (hCA, EC 4.2.1.1), hCA VI, has been cloned, expressed, and purified in a bacterial expression system. The kinetic parameters for the CO2 hydration reaction proved hCA VI to possess a kcat of 3.4 × 105 s−1 and kcat/KM of 4.9 × 107 M−1 s−1 (at pH 7.5 and 20 °C). hCA VI has a significant catalytic activity for the physiological reaction, of the same order of magnitude as the ubiquitous isoform CA I or the transmembrane, tumor-associated isozyme CA IX. A series of amino acids and amines were shown to act as CA VI activators, with variable efficacies. l-His, l-Trp, and dopamine showed weak CA VI activating effects (KAs in the range of 21–42 μM), whereas d-His, d-Phe, l-DOPA, l-Trp, serotonin, and some pyridyl-alkylamines were better activators, with KAs in the range of 13–19 μM. The best CA VI activators were l-Phe, d-DOPA, l-Tyr, 4-amino-l-Phe, and histamine, with KAs in the range of 1.23–9.31 μM. All these activators enhance kcat, having no effect on KM, participating thus in the rate determining step in the catalytic cycle, the proton transfer reactions between the enzyme active site and the environment.