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Claudiu T Supuran - One of the best experts on this subject based on the ideXlab platform.

  • Anion inhibition studies of a beta carbonic anhydrase from the malaria mosquito Anopheles gambiae
    Taylor & Francis Group, 2018
    Co-Authors: Daniela Vullo, Leo Syrjänen, Marianne Kuuslahti, Seppo Parkkila, Claudiu T Supuran
    Abstract:

    An anion inhibition study of the β-class carbonic anhydrase, AgaCA, from the malaria mosquito Anopheles gambiae is reported. A series of simple as well as complex inorganic anions, and small molecules known to interact with CAs were included in the study. Bromide, iodide, bisulphite, perchlorate, perrhenate, perruthenate, and peroxydisulphate were ineffective AgaCA inhibitors, with KIs > 200 mM. Fluoride, chloride, cyanate, thiocyanate, cyanide, bicarbonate, carbonate, nitrite, nitrate, sulphate, stannate, selenate, tellurate, diphosphate, divanadate, tetraborate, selenocyanide, and trithiocarbonate showed KIs in the range of 1.80–9.46 mM, whereas N,N-diethyldithiocarbamate was a submillimolar AgaCA inhibitor (KI of 0.65 mM). The most effective AgaCA inhibitors were sulphamide, sulphamic Acid, phenylboronic Acid and Phenylarsonic Acid, with inhibition constants in the range of 21–84 µM. The control of insect vectors responsible of the transmission of many protozoan diseases is rather difficult nowadays, and finding agents which can interfere with these processes, as the enzyme inhibitors investigated here, may arrest the spread of these diseases worldwide

  • Cloning, Characterization and Anion Inhibition Studies of a β-Carbonic Anhydrase from the Pathogenic Protozoan Entamoeba histolytica
    MDPI AG, 2018
    Co-Authors: Susanna Haapanen, Seppo Parkkila, Marianne Kuuslahti, Silvia Bua, Claudiu T Supuran
    Abstract:

    We report the cloning and catalytic activity of a β-carbonic anhydrase (CA, EC 4.2.1.1), isolated from the pathogenic protozoan Entamoeba histolytica, EhiCA. This enzyme has a high catalytic activity for the physiologic CO2 hydration reaction, with a kcat of 6.7 × 105 s−1 and a kcat/Km of 8.9 × 107 M−1 × s−1. An anion inhibition study of EhiCA with inorganic/organic anions and small molecules revealed that fluoride, chloride, cyanide, azide, pyrodiphosphate, perchlorate, tetrafluoroborate and sulfamic Acid did not inhibit the enzyme activity, whereas pseudohalides (cyanate and thiocyanate), bicarbonate, nitrate, nitrite, diethyldithiocarbamate, and many complex inorganic anions showed inhibition in the millimolar range (KIs of 0.51–8.4 mM). The best EhiCA inhibitors were fluorosulfonate, sulfamide, phenylboronic Acid and Phenylarsonic Acid (KIs in the range of 28–86 μM). Since β-CAs are not present in vertebrates, the present study may be useful for detecting lead compounds for the design of effective enzyme inhibitors, with potential to develop anti-infectives with alternative mechanisms of action

  • Sequence Analysis, Kinetic Constants, and Anion Inhibition Profile of the Nacrein-Like Protein (CgiNAP2X1) from the Pacific Oyster Magallana gigas (Ex-Crassostrea gigas)
    MDPI AG, 2017
    Co-Authors: Rosa Perfetto, Daniela Vullo, Sonia Del Prete, Claudiu T Supuran, Mose Rossi, Giovanni Sansone, Carmela M. A. Barone, Clemente Capasso
    Abstract:

    The carbonic anhydrase (CA, EC 4.2.1.1) superfamily of metalloenzymes catalyzes the hydration of carbon dioxide to bicarbonate and protons. The catalytically active form of these enzymes incorporates a metal hydroxide derivative, the formation of which is the rate-determining step of catalytic reaction, being affected by the transfer of a proton from a metal-coordinated water molecule to the environment. Here, we report the cloning, expression, and purification of a particular CA, i.e., nacrein-like protein encoded in the genome of the Pacific oyster Magallana gigas (previously known as Crassostrea gigas). Furthermore, the amino Acid sequence, kinetic constants, and anion inhibition profile of the recombinant enzyme were investigated for the first time. The new protein, CgiNAP2X1, is highly effective as catalyst for the CO2 hydration reaction, based on the measured kinetic parameters, i.e., kcat = 1.0 × 106 s−1 and kcat/KM = 1.2 × 108 M−1·s−1. CgiNAP2X1 has a putative signal peptide, which probably allows an extracellular localization of the protein. The inhibition data demonstrated that the best anion inhibitors of CgiNAP2X1 were diethyldithiocarbamate, sulfamide, sulfamate, phenylboronic Acid and Phenylarsonic Acid, which showed a micromolar affinity for this enzyme, with KIs in the range of 76–87 μM. These studies may add new information on the physiological role of the molluskan CAs in the biocalcification processes

  • Legionella pneumophila Carbonic Anhydrases: Underexplored Antibacterial Drug Targets
    MDPI AG, 2016
    Co-Authors: Claudiu T Supuran
    Abstract:

    Carbonic anhydrases (CAs, EC 4.2.1.1) are metalloenzymes which catalyze the hydration of carbon dioxide to bicarbonate and protons. Many pathogenic bacteria encode such enzymes belonging to the α-, β-, and/or γ-CA families. In the last decade, enzymes from some of these pathogens, including Legionella pneumophila, have been cloned and characterized in detail. These enzymes were shown to be efficient catalysts for CO2 hydration, with kcat values in the range of (3.4–8.3) × 105 s−1 and kcat/KM values of (4.7–8.5) × 107 M−1·s−1. In vitro inhibition studies with various classes of inhibitors, such as anions, sulfonamides and sulfamates, were also reported for the two β-CAs from this pathogen, LpCA1 and LpCA2. Inorganic anions were millimolar inhibitors, whereas diethyldithiocarbamate, sulfamate, sulfamide, phenylboronic Acid, and Phenylarsonic Acid were micromolar ones. The best LpCA1 inhibitors were aminobenzolamide and structurally similar sulfonylated aromatic sulfonamides, as well as acetazolamide and ethoxzolamide (KIs in the range of 40.3–90.5 nM). The best LpCA2 inhibitors belonged to the same class of sulfonylated sulfonamides, together with acetazolamide, methazolamide, and dichlorophenamide (KIs in the range of 25.2–88.5 nM). Considering such preliminary results, the two bacterial CAs from this pathogen represent promising yet underexplored targets for obtaining antibacterials devoid of the resistance problems common to most of the clinically used antibiotics, but further studies are needed to validate them in vivo as drug targets

  • anion inhibition studies of the dandruff producing fungus malassezia globosa β carbonic anhydrase mgca
    Bioorganic & Medicinal Chemistry Letters, 2015
    Co-Authors: Sonia Del Prete, Daniela Vullo, Clemente Capasso, Sameh M. Osman, Zeid A. Alothman, Claudiu T Supuran
    Abstract:

    The genome of the fungal parasite Malassezia globosa, the causative agent of dandruff, contains a single gene annotated as encoding a carbonic anhydrase (CAs, EC 4.2.1.1) belonging to the β-class (MgCA). In an earlier work (J. Med. Chem. 2012, 55, 3513) we have validated this enzyme as an anti-dandruff drug target, reporting that sulfonamide inhibitors show in vitro and in vivo effects, in an animal model of Malassezia infection. However, few classes of compounds apart the sulfonamides, were investigated for their activity against MgCA. Here we present an anion inhibition study of this enzyme, reporting that metal complexing anions such as cyanate, thiocyanate, cyanide, azide are weak MgCA inhibitors (KIs ranging between 6.81 and 45.2 mM) whereas bicarbonate (KI of 0.59 mM) and diethyldithiocarbamate (KI of 0.30 mM) together with sulfamide, sulfamate, phenylboronic Acid and Phenylarsonic Acid were the most effective inhibitors detected so far, with KIs ranging between 83 and 94 μM. This study may help a better understanding of the inhibition profile of this enzyme and may offer the possibility to design new such modulators of activity belonging to different chemical classes.

Toshikazu Kaise - One of the best experts on this subject based on the ideXlab platform.

  • determination of degradation compounds derived from lewisite by high performance liquid chromatography inductively coupled plasma mass spectrometry
    Applied Organometallic Chemistry, 2006
    Co-Authors: Kenji Kinoshita, Ozamu Shikino, Yasuo Seto, Toshikazu Kaise
    Abstract:

    A method for determination of 2-chlorovinylarsenous Acid (CVAA) and 2-chlorovinylarsonic Acid (CVAOA), which are degradation compounds of the chemical warfare agent Lewisite, was examined by high-performance liquid chromatography/inductively coupled plasma-mass spectrometry (HPLC/ICP-MS). Inertsil C 8 was suitable as the column and the mobile phase was consisted of 0.1% formic Acid-acetonitrile (80:20). These compounds were detected sensitively in a short time and separated from inorganic arsenicals and diphenylarsinic Acid (DPAA) and Phenylarsonic Acid (PAA), which are degradation compounds of diphenylchloroarsine and phenyldichloroarsine, respectively. The detection limits of CVAA and CVAOA were 0.2 and 0.1 ngAs/ml, respectively. In addition, a dynamic reaction cell and oxygen as the reaction gas were applied, and then arsenic was detected as AsO + (m/z 91) in order to prevent interference by ArCl + (m/z 75). This method was applied to the analysis of urine obtained from a CVAA-administered mouse and CVAOA was detected as the main metabolite. Thus, the speciation analysis of arsenic compounds derived from chemical warfare agents was achieved by HPLC/ICP-MS.

  • determination of diphenylarsinic Acid and Phenylarsonic Acid the degradation products of organoarsenic chemical warfare agents in well water by hplc icp ms
    Applied Organometallic Chemistry, 2005
    Co-Authors: Kenji Kinoshita, Masatoshi Morita, Yasuo Shida, Chiseko Sakuma, Mutsuo Ishizaki, Koichi Kiso, Osamu Shikino, Hiroyasu Ito, Takafumi Ochi, Toshikazu Kaise
    Abstract:

    Diphenylarsinic Acid (DPAA) and Phenylarsonic Acid (PAA), which were degradation products of organoarsenic chemical warfare agents used as sternutatory gas, were detected in the well water at Kamisu, Ibaraki Prefecture, Japan. The standard material of DPAA was synthesized with aqueous arsenic Acid and phenylhydrazine in order to determine organic arsenic compounds in well water. The DPAA showed a protonated ion at m/z 263 [M + H]+ and a loss of H2O ion at m/z 245 [M + H − H2O]+ from protonated ion by the electrospray ionization time-of-flight mass spectrometry. The quantitative analysis of DPAA and PAA was performed by high-performance liquid chromatography inductively coupled plasma mass spectrometry and the system worked well for limpid liquid samples such as well water. Copyright © 2005 John Wiley & Sons, Ltd.

Clemente Capasso - One of the best experts on this subject based on the ideXlab platform.

  • Sequence Analysis, Kinetic Constants, and Anion Inhibition Profile of the Nacrein-Like Protein (CgiNAP2X1) from the Pacific Oyster Magallana gigas (Ex-Crassostrea gigas)
    MDPI AG, 2017
    Co-Authors: Rosa Perfetto, Daniela Vullo, Sonia Del Prete, Claudiu T Supuran, Mose Rossi, Giovanni Sansone, Carmela M. A. Barone, Clemente Capasso
    Abstract:

    The carbonic anhydrase (CA, EC 4.2.1.1) superfamily of metalloenzymes catalyzes the hydration of carbon dioxide to bicarbonate and protons. The catalytically active form of these enzymes incorporates a metal hydroxide derivative, the formation of which is the rate-determining step of catalytic reaction, being affected by the transfer of a proton from a metal-coordinated water molecule to the environment. Here, we report the cloning, expression, and purification of a particular CA, i.e., nacrein-like protein encoded in the genome of the Pacific oyster Magallana gigas (previously known as Crassostrea gigas). Furthermore, the amino Acid sequence, kinetic constants, and anion inhibition profile of the recombinant enzyme were investigated for the first time. The new protein, CgiNAP2X1, is highly effective as catalyst for the CO2 hydration reaction, based on the measured kinetic parameters, i.e., kcat = 1.0 × 106 s−1 and kcat/KM = 1.2 × 108 M−1·s−1. CgiNAP2X1 has a putative signal peptide, which probably allows an extracellular localization of the protein. The inhibition data demonstrated that the best anion inhibitors of CgiNAP2X1 were diethyldithiocarbamate, sulfamide, sulfamate, phenylboronic Acid and Phenylarsonic Acid, which showed a micromolar affinity for this enzyme, with KIs in the range of 76–87 μM. These studies may add new information on the physiological role of the molluskan CAs in the biocalcification processes

  • anion inhibition studies of the dandruff producing fungus malassezia globosa β carbonic anhydrase mgca
    Bioorganic & Medicinal Chemistry Letters, 2015
    Co-Authors: Sonia Del Prete, Daniela Vullo, Clemente Capasso, Sameh M. Osman, Zeid A. Alothman, Claudiu T Supuran
    Abstract:

    The genome of the fungal parasite Malassezia globosa, the causative agent of dandruff, contains a single gene annotated as encoding a carbonic anhydrase (CAs, EC 4.2.1.1) belonging to the β-class (MgCA). In an earlier work (J. Med. Chem. 2012, 55, 3513) we have validated this enzyme as an anti-dandruff drug target, reporting that sulfonamide inhibitors show in vitro and in vivo effects, in an animal model of Malassezia infection. However, few classes of compounds apart the sulfonamides, were investigated for their activity against MgCA. Here we present an anion inhibition study of this enzyme, reporting that metal complexing anions such as cyanate, thiocyanate, cyanide, azide are weak MgCA inhibitors (KIs ranging between 6.81 and 45.2 mM) whereas bicarbonate (KI of 0.59 mM) and diethyldithiocarbamate (KI of 0.30 mM) together with sulfamide, sulfamate, phenylboronic Acid and Phenylarsonic Acid were the most effective inhibitors detected so far, with KIs ranging between 83 and 94 μM. This study may help a better understanding of the inhibition profile of this enzyme and may offer the possibility to design new such modulators of activity belonging to different chemical classes.

  • Cloning, characterization and anion inhibition studies of a new γ-carbonic anhydrase from the Antarctic bacterium Pseudoalteromonas haloplanktis.
    Bioorganic & medicinal chemistry, 2015
    Co-Authors: Viviana De Luca, Daniela Vullo, Sonia Del Prete, Vincenzo Carginale, Andrea Scozzafava, Sameh M. Osman, Zeid A. Alothman, Clemente Capasso
    Abstract:

    A new γ-class carbonic anhydrase (CA, EC 4.2.1.1) was cloned, purified and characterized from the Antarctic bacterium Pseudoalteromonas haloplanktis , PhaCAγ. The enzyme has a medium-low catalytic activity for the physiologic reaction of CO 2 hydration to bicarbonate and protons, with a k cat of 1.4 × 10 5  s −1 and a k cat / K m of 1.9 × 10 6  M −1  s −1 . An anion inhibition study of PhaCAγ with inorganic anions and small molecule inhibitors is also reported. Many anions present in sea water, such as chloride, fluoride, sulfate, iodide, but also others such as azide, perchlorate and tetrafluoroborate did not inhibit this enzyme. Pseudohalides such as cyanate, thiocyanate, cyanide, selenocyanide, and also bicarbonate, nitrate, nitrite and many complex inorganic anions showed inhibition in the millimolar range ( K I in the range of 1.7–9.3 mM). The best PhaCAγ inhibitors detected in this study were diethyldithiocarbamate ( K I of 0.96 mM) as well as sulfamide, sulfamate, phenylboronic Acid and Phenylarsonic Acid ( K I in the range of 82–91 μM). Since γ-CAs are poorly understood at this moment, being present in carboxysomes and thus involved in photosynthesis, this study may be relevant for a better understanding of these processes in Antarctic bacteria/cyanobacteria.

  • cloning characterization and anion inhibition study of the δ class carbonic anhydrase tweca from the marine diatom thalassiosira weissflogii
    Bioorganic & Medicinal Chemistry, 2014
    Co-Authors: Sonia Del Prete, Daniela Vullo, Andrea Scozzafava, Clemente Capasso, Claudiu T Supuran
    Abstract:

    Abstract We investigated the catalytic activity and inhibition of the δ-class carbonic anhydrase (CA, EC 4.2.1.1) from the marine diatom Thalassiosira weissflogii , TweCA. The enzyme, obtained by cloning the synthetic gene, was an efficient catalyst for the CO 2 hydration, its physiological reaction, with a k cat of 1.3 × 10 5  s −1 and a k cat / K M of 3.3 × 10 7  M −1  s −1 . A range of inorganic anions and small molecules were investigated as inhibitors of TweCA. Chloride and sulfate did not inhibit the enzyme ( K I s >200 mM) whereas other halides and pseudohalides were submillimolar–millimolar inhibitors ( K I s in the range of 0.93–8.3 mM). The best TweCA inhibitors were hydrogen sulfide, sulfamate, sulfamide, phenylboronic Acid and Phenylarsonic Acid, with K I s in the range of 9–90 μM, whereas acetazolamide inhibited the enzyme with a K I of 83 nM. This is the first kinetic and inhibition study of a δ-class CA. However, these enzymes are widespread in the marine phytoplankton, being present in haptophytes, dinoflagellates, diatoms, and chlorophytic prasinophytes, contributing to the CO 2 fixation by sea organisms. A phylogenetic analysis with all five genetic families of CAs showed that α- and δ-CAs are evolutionarily more related to each other with respect to the γ-CAs, although these three families clustered all together. On the contrary, the β- and ζ-CAs are also related to each other but phylogenetically much more distant from the α-, γ and δ-CA cluster. Thus, the study of δ-CAs is essential for better understanding this superfamily of metalloenzymes and their potential biotechnological applications in biomimetic CO 2 capture processes, as these enzymes are part of the carbon concentrating mechanism used by many photosynthetic organisms.

  • kinetic and anion inhibition studies of a β carbonic anhydrase fbica 1 from the c4 plant flaveria bidentis
    Bioorganic & Medicinal Chemistry Letters, 2013
    Co-Authors: Simona Maria Monti, Daniela Vullo, Andrea Scozzafava, Clemente Capasso, Giuseppina De Simone, Nina A Dathan, Martha Ludwig, Claudiu T Supuran
    Abstract:

    Abstract Several β-carbonic anhydrases (CAs, EC 4.2.1.1) are present in all land plants examined thus far. Here we report the first detailed biochemical characterization of one such isoform, FbiCA 1, from the C4 plant Flaveria bidentis, which was cloned, purified and characterized as recombinant protein. FbiCA 1 has an interesting CO2 hydrase catalytic activity (kcat of 1.2 × 105 and kcat/Km of 7.5 × 106 M−1 × s−1) and was moderately inhibited by most simple/complex inorganic anions. Potent FbiCA 1 inhibitors were also detected, such as trithiocarbonate, diethyldithiocarbamate, sulfamide, sulfamic Acid, phenylboronic Acid and Phenylarsonic Acid (KIs in the range of 4–60 μM). Such inhibitors may be used as tools to better understand the role of various β-CA isoforms in photosynthesis.

Kenji Kinoshita - One of the best experts on this subject based on the ideXlab platform.

  • determination of degradation compounds derived from lewisite by high performance liquid chromatography inductively coupled plasma mass spectrometry
    Applied Organometallic Chemistry, 2006
    Co-Authors: Kenji Kinoshita, Ozamu Shikino, Yasuo Seto, Toshikazu Kaise
    Abstract:

    A method for determination of 2-chlorovinylarsenous Acid (CVAA) and 2-chlorovinylarsonic Acid (CVAOA), which are degradation compounds of the chemical warfare agent Lewisite, was examined by high-performance liquid chromatography/inductively coupled plasma-mass spectrometry (HPLC/ICP-MS). Inertsil C 8 was suitable as the column and the mobile phase was consisted of 0.1% formic Acid-acetonitrile (80:20). These compounds were detected sensitively in a short time and separated from inorganic arsenicals and diphenylarsinic Acid (DPAA) and Phenylarsonic Acid (PAA), which are degradation compounds of diphenylchloroarsine and phenyldichloroarsine, respectively. The detection limits of CVAA and CVAOA were 0.2 and 0.1 ngAs/ml, respectively. In addition, a dynamic reaction cell and oxygen as the reaction gas were applied, and then arsenic was detected as AsO + (m/z 91) in order to prevent interference by ArCl + (m/z 75). This method was applied to the analysis of urine obtained from a CVAA-administered mouse and CVAOA was detected as the main metabolite. Thus, the speciation analysis of arsenic compounds derived from chemical warfare agents was achieved by HPLC/ICP-MS.

  • determination of diphenylarsinic Acid and Phenylarsonic Acid the degradation products of organoarsenic chemical warfare agents in well water by hplc icp ms
    Applied Organometallic Chemistry, 2005
    Co-Authors: Kenji Kinoshita, Masatoshi Morita, Yasuo Shida, Chiseko Sakuma, Mutsuo Ishizaki, Koichi Kiso, Osamu Shikino, Hiroyasu Ito, Takafumi Ochi, Toshikazu Kaise
    Abstract:

    Diphenylarsinic Acid (DPAA) and Phenylarsonic Acid (PAA), which were degradation products of organoarsenic chemical warfare agents used as sternutatory gas, were detected in the well water at Kamisu, Ibaraki Prefecture, Japan. The standard material of DPAA was synthesized with aqueous arsenic Acid and phenylhydrazine in order to determine organic arsenic compounds in well water. The DPAA showed a protonated ion at m/z 263 [M + H]+ and a loss of H2O ion at m/z 245 [M + H − H2O]+ from protonated ion by the electrospray ionization time-of-flight mass spectrometry. The quantitative analysis of DPAA and PAA was performed by high-performance liquid chromatography inductively coupled plasma mass spectrometry and the system worked well for limpid liquid samples such as well water. Copyright © 2005 John Wiley & Sons, Ltd.

Daniela Vullo - One of the best experts on this subject based on the ideXlab platform.

  • Anion inhibition studies of a beta carbonic anhydrase from the malaria mosquito Anopheles gambiae
    Taylor & Francis Group, 2018
    Co-Authors: Daniela Vullo, Leo Syrjänen, Marianne Kuuslahti, Seppo Parkkila, Claudiu T Supuran
    Abstract:

    An anion inhibition study of the β-class carbonic anhydrase, AgaCA, from the malaria mosquito Anopheles gambiae is reported. A series of simple as well as complex inorganic anions, and small molecules known to interact with CAs were included in the study. Bromide, iodide, bisulphite, perchlorate, perrhenate, perruthenate, and peroxydisulphate were ineffective AgaCA inhibitors, with KIs > 200 mM. Fluoride, chloride, cyanate, thiocyanate, cyanide, bicarbonate, carbonate, nitrite, nitrate, sulphate, stannate, selenate, tellurate, diphosphate, divanadate, tetraborate, selenocyanide, and trithiocarbonate showed KIs in the range of 1.80–9.46 mM, whereas N,N-diethyldithiocarbamate was a submillimolar AgaCA inhibitor (KI of 0.65 mM). The most effective AgaCA inhibitors were sulphamide, sulphamic Acid, phenylboronic Acid and Phenylarsonic Acid, with inhibition constants in the range of 21–84 µM. The control of insect vectors responsible of the transmission of many protozoan diseases is rather difficult nowadays, and finding agents which can interfere with these processes, as the enzyme inhibitors investigated here, may arrest the spread of these diseases worldwide

  • Sequence Analysis, Kinetic Constants, and Anion Inhibition Profile of the Nacrein-Like Protein (CgiNAP2X1) from the Pacific Oyster Magallana gigas (Ex-Crassostrea gigas)
    MDPI AG, 2017
    Co-Authors: Rosa Perfetto, Daniela Vullo, Sonia Del Prete, Claudiu T Supuran, Mose Rossi, Giovanni Sansone, Carmela M. A. Barone, Clemente Capasso
    Abstract:

    The carbonic anhydrase (CA, EC 4.2.1.1) superfamily of metalloenzymes catalyzes the hydration of carbon dioxide to bicarbonate and protons. The catalytically active form of these enzymes incorporates a metal hydroxide derivative, the formation of which is the rate-determining step of catalytic reaction, being affected by the transfer of a proton from a metal-coordinated water molecule to the environment. Here, we report the cloning, expression, and purification of a particular CA, i.e., nacrein-like protein encoded in the genome of the Pacific oyster Magallana gigas (previously known as Crassostrea gigas). Furthermore, the amino Acid sequence, kinetic constants, and anion inhibition profile of the recombinant enzyme were investigated for the first time. The new protein, CgiNAP2X1, is highly effective as catalyst for the CO2 hydration reaction, based on the measured kinetic parameters, i.e., kcat = 1.0 × 106 s−1 and kcat/KM = 1.2 × 108 M−1·s−1. CgiNAP2X1 has a putative signal peptide, which probably allows an extracellular localization of the protein. The inhibition data demonstrated that the best anion inhibitors of CgiNAP2X1 were diethyldithiocarbamate, sulfamide, sulfamate, phenylboronic Acid and Phenylarsonic Acid, which showed a micromolar affinity for this enzyme, with KIs in the range of 76–87 μM. These studies may add new information on the physiological role of the molluskan CAs in the biocalcification processes

  • anion inhibition studies of the dandruff producing fungus malassezia globosa β carbonic anhydrase mgca
    Bioorganic & Medicinal Chemistry Letters, 2015
    Co-Authors: Sonia Del Prete, Daniela Vullo, Clemente Capasso, Sameh M. Osman, Zeid A. Alothman, Claudiu T Supuran
    Abstract:

    The genome of the fungal parasite Malassezia globosa, the causative agent of dandruff, contains a single gene annotated as encoding a carbonic anhydrase (CAs, EC 4.2.1.1) belonging to the β-class (MgCA). In an earlier work (J. Med. Chem. 2012, 55, 3513) we have validated this enzyme as an anti-dandruff drug target, reporting that sulfonamide inhibitors show in vitro and in vivo effects, in an animal model of Malassezia infection. However, few classes of compounds apart the sulfonamides, were investigated for their activity against MgCA. Here we present an anion inhibition study of this enzyme, reporting that metal complexing anions such as cyanate, thiocyanate, cyanide, azide are weak MgCA inhibitors (KIs ranging between 6.81 and 45.2 mM) whereas bicarbonate (KI of 0.59 mM) and diethyldithiocarbamate (KI of 0.30 mM) together with sulfamide, sulfamate, phenylboronic Acid and Phenylarsonic Acid were the most effective inhibitors detected so far, with KIs ranging between 83 and 94 μM. This study may help a better understanding of the inhibition profile of this enzyme and may offer the possibility to design new such modulators of activity belonging to different chemical classes.

  • Cloning, characterization and anion inhibition studies of a new γ-carbonic anhydrase from the Antarctic bacterium Pseudoalteromonas haloplanktis.
    Bioorganic & medicinal chemistry, 2015
    Co-Authors: Viviana De Luca, Daniela Vullo, Sonia Del Prete, Vincenzo Carginale, Andrea Scozzafava, Sameh M. Osman, Zeid A. Alothman, Clemente Capasso
    Abstract:

    A new γ-class carbonic anhydrase (CA, EC 4.2.1.1) was cloned, purified and characterized from the Antarctic bacterium Pseudoalteromonas haloplanktis , PhaCAγ. The enzyme has a medium-low catalytic activity for the physiologic reaction of CO 2 hydration to bicarbonate and protons, with a k cat of 1.4 × 10 5  s −1 and a k cat / K m of 1.9 × 10 6  M −1  s −1 . An anion inhibition study of PhaCAγ with inorganic anions and small molecule inhibitors is also reported. Many anions present in sea water, such as chloride, fluoride, sulfate, iodide, but also others such as azide, perchlorate and tetrafluoroborate did not inhibit this enzyme. Pseudohalides such as cyanate, thiocyanate, cyanide, selenocyanide, and also bicarbonate, nitrate, nitrite and many complex inorganic anions showed inhibition in the millimolar range ( K I in the range of 1.7–9.3 mM). The best PhaCAγ inhibitors detected in this study were diethyldithiocarbamate ( K I of 0.96 mM) as well as sulfamide, sulfamate, phenylboronic Acid and Phenylarsonic Acid ( K I in the range of 82–91 μM). Since γ-CAs are poorly understood at this moment, being present in carboxysomes and thus involved in photosynthesis, this study may be relevant for a better understanding of these processes in Antarctic bacteria/cyanobacteria.

  • cloning characterization and anion inhibition study of the δ class carbonic anhydrase tweca from the marine diatom thalassiosira weissflogii
    Bioorganic & Medicinal Chemistry, 2014
    Co-Authors: Sonia Del Prete, Daniela Vullo, Andrea Scozzafava, Clemente Capasso, Claudiu T Supuran
    Abstract:

    Abstract We investigated the catalytic activity and inhibition of the δ-class carbonic anhydrase (CA, EC 4.2.1.1) from the marine diatom Thalassiosira weissflogii , TweCA. The enzyme, obtained by cloning the synthetic gene, was an efficient catalyst for the CO 2 hydration, its physiological reaction, with a k cat of 1.3 × 10 5  s −1 and a k cat / K M of 3.3 × 10 7  M −1  s −1 . A range of inorganic anions and small molecules were investigated as inhibitors of TweCA. Chloride and sulfate did not inhibit the enzyme ( K I s >200 mM) whereas other halides and pseudohalides were submillimolar–millimolar inhibitors ( K I s in the range of 0.93–8.3 mM). The best TweCA inhibitors were hydrogen sulfide, sulfamate, sulfamide, phenylboronic Acid and Phenylarsonic Acid, with K I s in the range of 9–90 μM, whereas acetazolamide inhibited the enzyme with a K I of 83 nM. This is the first kinetic and inhibition study of a δ-class CA. However, these enzymes are widespread in the marine phytoplankton, being present in haptophytes, dinoflagellates, diatoms, and chlorophytic prasinophytes, contributing to the CO 2 fixation by sea organisms. A phylogenetic analysis with all five genetic families of CAs showed that α- and δ-CAs are evolutionarily more related to each other with respect to the γ-CAs, although these three families clustered all together. On the contrary, the β- and ζ-CAs are also related to each other but phylogenetically much more distant from the α-, γ and δ-CA cluster. Thus, the study of δ-CAs is essential for better understanding this superfamily of metalloenzymes and their potential biotechnological applications in biomimetic CO 2 capture processes, as these enzymes are part of the carbon concentrating mechanism used by many photosynthetic organisms.