title: Carbonic anhydrase activators: the first activation study of the human secretory isoform VI with amino acids and amines.
abstract: 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 CO(2) hydration reaction proved hCA VI to possess a k(cat) of 3.4 x 10(5)s(-1) and k(cat)/K(M) of 4.9 x 10(7)M(-1)s(-1) (at pH 7.5 and 20 degrees 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 (K(A)s in the range of 21-42 microM), whereas d-His, d-Phe, l-DOPA, l-Trp, serotonin, and some pyridyl-alkylamines were better activators, with K(A)s in the range of 13-19 microM. The best CA VI activators were l-Phe, d-DOPA, l-Tyr, 4-amino-l-Phe, and histamine, with K(A)s in the range of 1.23-9.31 microM. All these activators enhance k(cat), having no effect on K(M), participating thus in the rate determining step in the catalytic cycle, the proton transfer reactions between the enzyme active site and the environment.
authors.affiliation: Department of Gastroenterology and Hepatology, Kochi Medical School, Nankoku, Kochi, Japan.
authors.forename: Isao
authors.initials: I
authors.lastname: Nishimori
authors.forename: Saburo
authors.initials: S
authors.lastname: Onishi
authors.forename: Daniela
authors.initials: D
authors.lastname: Vullo
authors.forename: Alessio
authors.initials: A
authors.lastname: Innocenti
authors.forename: Andrea
authors.initials: A
authors.lastname: Scozzafava
authors.forename: Claudiu T
authors.initials: CT
authors.lastname: Supuran
mesh_terms: D000588:Amines / Q000494:pharmacology*; D000595:Amino Acid Sequence; D000596:Amino Acids / Q000494:pharmacology*; D002256:Carbonic Anhydrases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D003001:Cloning, Molecular; D004789:Enzyme Activation / Q000187:drug effects; D015971:Gene Expression Regulation, Enzymologic; D006801:Humans; D008969:Molecular Sequence Data; D020033:Protein Isoforms / Q000819:agonists / Q000235:genetics / Q000378:metabolism; D021122:Protein Subunits / Q000819:agonists* / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
chemical_list: D000588:Amines; D000596:Amino Acids; D020033:Protein Isoforms; D021122:Protein Subunits; D002256:Carbonic Anhydrases; C477051:carbonic anhydrase VI