Abstract
Background
Carbonic anhydrase (CA) catalyzes the reversible hydration of CO2 to bicarbonate and a proton, and α-class CA has been reported to facilitate the acid acclimation of Helicobacter pylori (hpαCA). The purpose of this study was to characterize the β-class CA of H. pylori (hpβCA) and elucidate the role of this enzyme as a possible drug target for eradication therapy.
Methods
We isolated DNA clones of independent H. pylori strains obtained from patients with gastritis (n = 15), gastric ulcer (n = 6), or gastric cancer (n = 16), and then studied genetic polymorphisms. In addition, the susceptibility of H. pylori to sulpiride, an antiulcer drug and efficient inhibitor of both hpαCA and hpβCA, was studied with an in vitro killing assay.
Results
DNA sequences of all 37 hpβCA clones encoded a 221 amino acid polypeptide with a variety of polymorphisms (57 types of amino acid substitution at 48 residue positions). There was no polymorphism functionally relevant to the gastric lesion type. One strain included unique residues that were not seen in the other 36 clones from Japanese patients but which were found in a strain obtained from the United Kingdom. Sulpiride had killing effects at concentrations greater than 200 μg/ml for H. pylori, including strains resistant to clarithromycin, metronidazole, or ampicillin.
Conclusions
Helicobacter pylori might have evolved independently in the Caucasian and Japanese populations. Dual inhibition of α-and β-class CAs could be applied as alternative therapy for eradication of H. pylori.
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References
Supuran CT, Scozzafava A, Conway J. Carbonic anhydrase: its inhibitors and activators. Boca Raton: CRC Press; 2004.
Sly WS, Hu PY. Human carbonic anhydrases and carbonic anhydrase deficiencies. Annu Rev Biochem 1995;64:375–401.
Chegwidden WR, Carter N. Introduction to the carbonic anhydrases. In: Chegwidden WR, Carter ND, Edwards YH, editors. The carbonic hydrases. New Horizons. Basel: Birkhäuser Verlag; 2000.p. 13–28.
Lehtonen J, Shen B, Vihinen M, Casini A, Scozzafava A, Supuran CT, et al. Characterization of CA XIII, a novel member of the carbonic anhydrase isozyme family. J Biol Chem 2004;279:2719–2727.
Fujikawa-Adachi K, Nishimori I, Taguchi T, Onishi S. Human carbonic anhydrase XIV (CA14): cDNA cloning, mRNA expression, and mapping to chromosome 1. Genomics 1999;61:74–81.
Fujikawa-Adachi K, Nishimori I, Taguchi T, Onishi S. Human mitochondrial carbonic anhydrase VB. cDNA cloning, mRNA expression, subcellular localization, and mapping to chromosome X. J Biol Chem 1999;274:21228–21233.
Smith KS, Jakubzick C, Whittam TS, Ferry JG. Carbonic anhydrase is an ancient enzyme widespread in prokaryotes. Proc Natl Acad Sci USA 1999;96:15184–15189.
Fujikawa-Adachi K, Nishimori I, Taguchi T, Yuri K, Onishi S. cDNA sequence, mRNA expression, and chromosomal localization of human carbonic anhydrase-related protein, CA-RP XI. Biochim Biophys Acta 1999;1431:518–524.
Okamoto N, Fujikawa-Adachi K, Nishimori I, Taniuchi K, Onishi S. cDNA sequence of human carbonic anhydrase-related protein, CA-RP X: mRNA expressions of CA-RP X and XI in human brain. Biochim Biophys Acta 2001;1518:311–316.
Nishimori I, Minakuchi T, Onishi S, Vullo D, Scozzafava A, Supuran CT. Carbonic anhydrase inhibitors. DNA cloning, characterization, and inhibition studies of the human secretory isoform VI, a new target for sulfonamide and sulfamate inhibitors. J Med Chem 2007;50:381–388.
Vullo D, Franchi M, Gallori E, Antel J, Scozzafava A, Supuran CT. Carbonic anhydrase inhibitors. Inhibition of mitochondrial isozyme V with aromatic and heterocyclic sulfonamides. J Med Chem 2004;47:1272–1279.
Nishimori I, Vullo D, Innocenti A, Scozzafava A, Mastrolorenzo A, Supuran CT. Carbonic anhydrase inhibitors. The mitochondrial isozyme VB as a new target for sulfonamide and sulfamate inhibitors. J Med Chem 2005;48:7860–7866.
Supuran CT. Indisulam. An anticancer sulfonamide in clinical development. Expert Opin Investig Drugs 2003;12:283–287.
Weber A, Casini A, Heine A, Kuhn D, Supuran CT, Scozzafava A, et al. Unexpected nanomolar inhibition of carbonic anhydrase by COX-2-selective celecoxib: new pharmacological opport unities due to related binding site recognition. J Med Chem 2004;47:550–557.
Pastorekova S, Casini A, Scozzafava A, Vullo D, Pastorek J, Supuran CT. Carbonic anhydrase inhibitors: the first selective, membrane-impermeant inhibitors targeting the tumor-associated isozyme IX. Bioorg Med Chem Lett 2004;14:869–873.
Vullo D, Voipio J, Innocenti A, Rivera C, Ranki H, Scozzafava A, et al. Carbonic anhydrase inhibitors. Inhibition of the human cytosolic isozyme VII with aromatic and heterocyclic sulfonamides. Bioorg Med Chem Lett 2005;15:971–976.
De Simone G, Di Fiore A, Menchise V, Pedone C, Antel J, Casini A, et al. Carbonic anhydrase inhibitors. Zonisamide is an effective inhibitor of the cytosolic isozyme II and mitochondrial isozyme V: solution and X-ray crystallographic studies. Bioorg Med Chem Lett 2005;15:2315–2320.
Vullo D, Innocenti A, Nishimori I, Pastorek J, Scozzafava A, Pastorekova S, et al. Carbonic anhydrase inhibitors. Inhibition of the transmembrane isozyme XII with sulfonamides-a new target for the design of antitumor and antiglaucoma drugs? Bioorg Med Chem Lett 2005;15:963–969.
Svastova E, Hulikova A, Rafajova M, Zat’ovicova M, Gibadulinova A, Casini A, et al. Hypoxia activates the capacity of tumor-associated carbonic anhydrase IX to acidify extracellular pH. FEBS Lett 2004;577:439–445.
Cecchi A, Hulikova A, Pastorek J, Pastorekova S, Scozzafava A, Winum JY, et al. Carbonic anhydrase inhibitors. Design of fluorescent sulfonamides as probes of tumor-associated carbonic anhydrase IX that inhibit isozyme IX-mediated acidification of hypoxic tumors. J Med Chem 2005;48:4834–4841.
Menchise V, De Simone G, Alterio V, Di Fiore A, Pedone C, Scozzafava A, et al. Carbonic anhydrase inhibitors: stacking with Phe131 determines active site binding region of inhibitors as exemplified by the X-ray crystal structure of a membrane-impermeant antitumor sulfonamide complexed with isozyme II. J Med Chem 2005;48:5721–5727.
Krungkrai J, Scozzafava A, Reungprapavut S, Krungkrai SR, Rattanajak R, Kamchonwongpaisan S, et al. Carbonic anhydrase inhibitors. Inhibition of Plasmodium falciparum carbonic anhydrase with aromatic sulfonamides: towards antimalarials with a novel mechanism of action? Bioorg Med Chem 2005;13:483–489.
Suarez Covarrubias A, Larsson AM, Hogbom M, Lindberg J, Bergfors T, Bjorkelid C, et al. Structure and function of carbonic anhydrases from Mycobacterium tuberculosis. J Biol Chem 2005;280:18782–18789.
Covarrubias AS, Bergfors T, Jones TA, Hogbom M. Structural mechanics of the pH-dependent activity of beta-carbonic anhydrase from Mycobacterium tuberculosis. J Biol Chem 2006;281:4993–4999.
Mogensen EG, Janbon G, Chaloupka J, Steegborn C, Fu MS, Moyrand F, et al. Cryptococcus neoformans senses CO2 through the carbonic anhydrase Can2 and the adenylyl cyclase Cac1. Eukaryot Cell 2006;5:103–111.
Klengel T, Liang WJ, Chaloupka J, Ruoff C, Schroppel K, Naglik JR, et al. Fungal adenylyl cyclase integrates CO2 sensing with cAMP signaling and virulence. Curr Biol 2005;15:2021–2026.
Suzuki H, Hibi T, Marshall BJ. Helicobacter pylori: present status and future prospects in Japan. J Gastroenterol 2007;42:1–15.
Malfertheiner P, Megraud F, O’Morain C, Hungin AP, Jones R, Axon A, et al. Current concepts in the management of Helicobacter pylori infection—the Maastricht 2-2000 Consensus Report. Aliment Pharmacol Ther 2002;16:167–180.
Lam SK, Talley NJ. Report of the 1997 Asia Pacific Consensus Conference on the management of Helicobacter pylori infection. J Gastroenterol Hepatol 1998;13:1–12.
Huang JQ, Hunt RH. Treatment after failure: the problem of ‘non-responders’. Gut 1 1999;45Suppl 1:I40–I44.
Laheij RJ, Rossum LG, Jansen JB, Straatman H, Verbeek AL. Evaluation of treatment regimens to cure Helicobacter pylori infection—a meta-analysis. Aliment Pharmacol Ther 1999;13:857–864.
Fischbach LA, van Zanten S, Dickason J. Meta-analysis: the efficacy, adverse events, and adherence related to first-line anti-Helicobacter pylori quadruple therapies. Aliment Pharmacol Ther 2004;20:1071–1082.
Gisbert JP, Pajares JM. Helicobacter pylori therapy: first-line options and rescue regimen. Dig Dis 2001;19:134–143.
Chi CH, Lin CY, Sheu BS, Yang HB, Huang AH, Wu JJ. Quadruple therapy containing amoxicillin and tetracycline is an effective regimen to rescue failed triple therapy by overcoming the antimicrobial resistance of Helicobacter pylori. Aliment Pharmacol Ther 2003;18:347–353.
Lamouliatte H, Megraud F, Delchier JC, Bretagne JF, Courillon-Mallet A, De Korwin JD, et al. Second-line treatment for failure to eradicate Helicobacter pylori: a randomized trial comparing four treatment strategies. Aliment Pharmacol Ther 2003;18:791–797.
Mantzaris GJ, Petraki K, Archavlis E, Amberiadis P, Christoforidis P, Kourtessas D, et al. Omeprazole triple therapy versus omeprazole quadruple therapy for healing duodenal ulcer and eradication of Helicobacter pylori infection: a 24-month follow-up study. Eur J Gastroenterol Hepatol 2002;14:1237–1243.
Sachs G, Weeks DL, Wen Y, Marcus EA, Scott DR, Melchers K. Acid acclimation by Helicobacter pylori. Physiology (Bethesda) 2005;20:429–438.
Marcus EA, Moshfegh AP, Sachs G, Scott DR. The periplasmic alpha-carbonic anhydrase activity of Helicobacter pylori is essential for acid acclimation. J Bacteriol 2005;187:729–738.
Stahler FN, Ganter L, Lederer K, Kist M, Bereswill S. Mutational analysis of the Helicobacter pylori carbonic anhydrases. FEMS Immunol Med Microbiol 2005;44:183–189.
Nishimori I, Minakuchi T, Morimoto K, Sano S, Onishi S, Takeuchi H, et al. Carbonic anhydrase inhibitors: DNA cloning and inhibition studies of the alpha-carbonic anhydrase from Helicobacter pylori, a new target for developing sulfonamide and sulfamate gastric drugs. J Med Chem 2006;49:2117–2126.
Nishimori I, Minakuchi T, Kohsaki T, Onishi S, Takeuchi H, Vullo D, et al. Carbonic anhydrase inhibitors: the beta-carbonic anhydrase from Helicobacter pylori is a new target for sulfonamide and sulfamate inhibitors. Bioorg Med Chem Lett 2007;17:3585–3594.
Tomb JF, White O, Kerlavage AR, Clayton RA, Sutton GG, Fleischmann RD, et al. The complete genome sequence of the gastric pathogen Helicobacter pylori. Nature 1997;388:539–547.
Bury-Moné S, Mendz GL, Ball GE, Thibonnier M, Stingl K, Ecobichon C, et al. Roles of alpha and beta carbonic anhydrases of Helicobacter pylori in the urease-dependent response to acidity and in colonization of the murine gastric mucosa. Infect Immun 2008;76:497–509.
Sano S, Nishimori I, Kosaki T, Fujikawa-Adachi K, Onishi S. The fine form of Helicobacter pylori on the metaplastic duodenal mucosa is associated with recurrent duodenal ulcers. Digestion 2001;64:161–168.
Takeuchi H, Shirai M, Akada JK, Tsuda M, Nakazawa T. Nucleotide sequence and characterization of cdrA, a cell division-related gene of Helicobacter pylori. J Bacteriol 1998;180:5263–5268.
Takeuchi H, Nakazawa T, Okamoto T, Shirai M, Kimoto M, Nishioka M, et al. Cell elongation and cell death of Helicobacter pylori is modulated by the disruption of cdrA (cell divisionrelated gene A). Microbiol Immunol 2006;50:487–497.
Cronk JD, Endrizzi JA, Cronk MR, O’neill JW, Zhang KY. Crystal structure of E. coli beta-carbonic anhydrase, an enzyme with an unusual pH-dependent activity. Protein Sci 2001;10:911–922.
Mitsuhashi S, Mizushima T, Yamashita E, Yamamoto M, Kumasaka T, Moriyama H, et al. X-ray structure of beta-carbonic anhydrase from the red alga, Porphyridium purpureum, reveals a novel catalytic site for CO(2) hydration. J Biol Chem 2000;275:5521–5526.
Hiltonen T, Bjorkbacka H, Forsman C, Clarke AK, Samuelsson G. Intracellular beta-carbonic anhydrase of the unicellular green alga Coccomyxa. Cloning of the cDNA and characterization of the functional enzyme overexpressed in Escherichia coli. Plant Physiol 1998;117:1341–1349.
Kimber MS, Pai EF. The active site architecture of Pisum sativum beta-carbonic anhydrase is a mirror image of that of alphacarbonic anhydrases. EMBO J 2000;19:1407–1418.
Chirica LC, Elleby B, Lindskog S. Cloning, expression and some properties of alpha-carbonic anhydrase from Helicobacter pylori. Biochim Biophys Acta 2001;1544:55–63.
Abbate F, Coetzee A, Casini A, Ciattini S, Scozzafava A, Supuran CT. Carbonic anhydrase inhibitors: X ray crystallographic structure of the adduct of human isozyme II with the antipsychotic drug sulpiride. Bioorg Med Chem Lett 2004;14:337–341.
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Morishita, S., Nishimori, I., Minakuchi, T. et al. Cloning, polymorphism, and inhibition of β-carbonic anhydrase of Helicobacter pylori . J Gastroenterol 43, 849–857 (2008). https://doi.org/10.1007/s00535-008-2240-3
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DOI: https://doi.org/10.1007/s00535-008-2240-3