title: Cloning, characterization and anion inhibition studies of a γ-carbonic anhydrase from the Antarctic bacterium Colwellia psychrerythraea.
abstract: We have cloned, purified and characterized the γ-carbonic anhydrase (CA, EC 4.2.1.1) present in the genome of the Antarctic bacterium Colwellia psychrerythraea, which is an obligate psychrophile. The enzyme shows a significant catalytic activity for the physiologic reaction of CO2 hydration to bicarbonate and protons, with the following kinetic parameters: kcat of 6.0×10(5)s(-1) and a kcat/Km of 4.7×10(6)M(-1)×s(-1). This activity was inhibited by the sulfonamide CA inhibitor (CAI) acetazolamide, with a KI of 502nM. A range of anions was also investigated for their inhibitory action against the new enzyme CpsCA. Perchlorate, tetrafluoroborate, fluoride and bromide were not inhibitory, whereas cyanate, thiocyanate, cyanide, hydrogensulfide, carbonate and bicarbonate showed KIs in the range of 1.4-4.4mM. Diethyldithiocarbamate was a better inhibitor (KI of 0.58mM) whereas sulfamide, sulfamate, phenylboronic acid and phenylarsonic acid were the most effective inhibitors detected, with KIs ranging between 8 and 38μM. The present study may shed some more light regarding the role that γ-CAs play in the life cycle of psychrophilic bacteria as the Antarctic one investigated here.
authors.affiliation: Istituto di Bioscienze e Biorisorse, CNR, Via Pietro Castellino 81, Napoli, Italy.
authors.forename: Viviana
authors.initials: V
authors.lastname: De Luca
authors.affiliation: Università degliStudi di Firenze, Dipartimento Di Chimica, Laboratorio di Chimica Bioinorganica, Polo Scientifico, Via della Lastruccia 3, 50019 Sesto Fiorentino, Florence, Italy.
authors.forename: Daniela
authors.initials: D
authors.lastname: Vullo
authors.affiliation: Istituto di Bioscienze e Biorisorse, CNR, Via Pietro Castellino 81, Napoli, Italy; Università degliStudi di Firenze, Dipartimento Di Chimica, Laboratorio di Chimica Bioinorganica, Polo Scientifico, Via della Lastruccia 3, 50019 Sesto Fiorentino, Florence, Italy.
authors.forename: Sonia
authors.initials: S
authors.lastname: Del Prete
authors.affiliation: Istituto di Bioscienze e Biorisorse, CNR, Via Pietro Castellino 81, Napoli, Italy.
authors.forename: Vincenzo
authors.initials: V
authors.lastname: Carginale
authors.affiliation: Department of Chemistry, College of Science, King Saud University, PO Box 2455, Riyadh 11451, Saudi Arabia; Advanced Materials Research Chair, Chemistry Department, College of Science, King Saud University, Riyadh 11451, Saudi Arabia.
authors.forename: Sameh M
authors.initials: SM
authors.lastname: Osman
authors.affiliation: Department of Chemistry, College of Science, King Saud University, PO Box 2455, Riyadh 11451, Saudi Arabia; Advanced Materials Research Chair, Chemistry Department, College of Science, King Saud University, Riyadh 11451, Saudi Arabia.
authors.forename: Zeid
authors.initials: Z
authors.lastname: AlOthman
authors.affiliation: Università degliStudi di Firenze, Dipartimento Di Chimica, Laboratorio di Chimica Bioinorganica, Polo Scientifico, Via della Lastruccia 3, 50019 Sesto Fiorentino, Florence, Italy; Department of Chemistry, College of Science, King Saud University, PO Box 2455, Riyadh 11451, Saudi Arabia; Università degli Studi di Firenze, Dipartimento Neurofarba, Sezione di Scienze Farmaceutiche, Polo Scientifico, Via U. Schiff 6, 50019 Sesto Fiorentino, Florence, Italy. Electronic address: claudiu.supuran@unifi.it.
authors.forename: Claudiu T
authors.initials: CT
authors.lastname: Supuran
authors.affiliation: Istituto di Bioscienze e Biorisorse, CNR, Via Pietro Castellino 81, Napoli, Italy. Electronic address: clemente.capasso@ibbr.cnr.it.
authors.forename: Clemente
authors.initials: C
authors.lastname: Capasso
mesh_terms: D000086:Acetazolamide / Q000494:pharmacology; D034241:Alteromonadaceae / Q000145:classification / Q000187:drug effects / Q000201:enzymology* / Q000235:genetics; D000595:Amino Acid Sequence; D000838:Anions / Q000494:pharmacology*; D000864:Antarctic Regions; D001426:Bacterial Proteins / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D002257:Carbonic Anhydrase Inhibitors / Q000494:pharmacology*; D002256:Carbonic Anhydrases / Q000235:genetics / Q000378:metabolism*; D003001:Cloning, Molecular; D057075:Enzyme Assays; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D007700:Kinetics; D008969:Molecular Sequence Data; D016366:Open Reading Frames; D010802:Phylogeny; D016415:Sequence Alignment
chemical_list: D000838:Anions; D001426:Bacterial Proteins; D002257:Carbonic Anhydrase Inhibitors; D002256:Carbonic Anhydrases; D000086:Acetazolamide
keywords: Anions; Antarctic carbonic anhydrase; Carbonic anhydrase; Cold adaptation; Cold-enzymes; Hydratase activity; Inhibitors; Metalloenzymes; Psychrophiles