Properties Of Carbonic Anhydrase Research Articles

Kinetic Properties of Carbonic Anhydrase Purified from Gills of Rainbow Trout (Oncorhynchus mykiss)

Hisar, O., Beydemir, Ş., Bülbül, M. and Yanik, T. 2006. Kinetic properties of carbonic anhydrase purified from gills of rainbow trout (Oncorhynchus mykiss). J. Appl. Anim. Res., 30: 185–188. Kinetic behavior and some properties of carbonic anhydrase enzyme purified from gills of rainbow trout (Oncorhynchus mykiss) were studied at 4C. The purification steps included high-speed centrifugation, sepharose-4B-L tyrosine-sulfanilamide affinity gel chromatography and dialysis. Yield and specific activity of the enzyme were 40% and 55.56 EU/mg protein, respectively. The overall purification was 104.8-fold. The molecular mass was approximately estimated as 28 kDa by SDS polyacrylamide gel electrophoresis and as 27 kDa by gel filtration column chromatography. Optimal pH, stable pH and optimal temperature of the enzyme were 9.5, 8.2 in 0.025 M boric acid buffer and 17.5C, respectively. KMand Vmax values of the enzyme for the substrate (p-nitrophenylacetate) were 8.13 mM and 2.10 μmol/mg protein/min, respectively. The dissociation constant of the enzyme inhibitor complex (Ki) value was 3.93±0.65 μM for sulfanilamide and sulfanilamide inhibited the enzyme in a noncompetitive manner. Data from present study showed that optimum kinetic properties of gill CA enzyme were different from other vertebrate CA's.

Purification and some properties of carbonic anhydrase from bovine skeletal muscle

Procedures for the purification of bovine muscle carbonic anhydrase (isoenzyme III) are described. The purified enzyme has a molecular weight near 29,000 and contains one Zn 2+ ion per molecule. The sedimentation coefficient, S 20,w 0, is 2.8 · 10 −13 s, the isoelectric pH is 8.5, and A 280 0.1% = 2.07 cm −1. The CO 2 hydration activity, expressed as k cat K m , is about 1.5% of that of human isoenzyme I (or B) and about 0.3% of that of human isoenzyme II (or C) at pH 8 and 25 °C. The activity is nearly independent of pH between pH 6.0 and 8.6. The muscle enzyme is weakly inhibited by the sulfonamide inhibitor, acetazolamide, whereas some anions, particularly sulfide and cyanate, are efficient inhibitors. Bovine carbonic anhydrase III contains five thiol groups, two of which react readily with Ellman's reagent without effect on the catalytic activity. A reinvestigation of the amino acid sequences of cysteine-containing tryptic peptides has shown that cysteine residues occur at sequence positions 66, 183, 188, 203, and 206.

Properties of Carbonic Anhydrase from the Saliva of the Rat

Properties of Carbonic Anhydrase from the Saliva of the Rat

Purification and properties of carbonic anhydrase from salmon erythrocytes

1. 1. Carbonic anhydrase (CA) from erythrocytes of the pink salmon, Onchorhyncus gorbushka, was purified using chloroform-ethanol extraction and Sephadex G-75 gel filtration. 2. 2. A single, high specific-activity CA isozyme having a molecular weight of 29,000 was found. The enzyme sedimented as a single boundary at a sedimentation velocity of 2.9S. 3. 3. Amino acid analysis revealed a composition similar to other submammalian CAs with the exception that the cysteine content was low (1 mol cysteine/mol enzyme). 4. 4. Like other submammalian CAs, the presence of a sulfhydryl reducing agent was required to maintain full activity and to prevent structural changes in the enzyme.

Purification and properties of carbonic anhydrase from Chlamydomonas reinhardii

Carbonic anhydrase (CA) was purified from the unicellular green alga Chlamydomonas reinhardii, and the purity of the preparation was established by gradient gel electrophoresis. The purified enzyme exhibited a MW of 165 000 and contained 6 atoms of Zn. The subunit MW, as determined by dodecyl sulfate electrophoresis, was 27 000. These results are consistent with a quarternary structure which is hexameric, each monomer containing 1 g atom of Zn. Like spinach CA, and in contrast to other oligomeric plant CAs, a sulfhydryl reducing agent is not needed to stabilize the enzyme. CO 2-hydrase activity was inhibited by both acetazolamide ( I 50 = 7.8 × 10 −9M) and sulfanilamide ( I 50 = 1.3 × 10 −5M), as well as by certain inorganic anions. The purified enzyme showed relatively weak esterase activity with p-nitrophenyl acetate but was an extremely effective esterase with 2-hydroxy-5-nitro-α-toluenesulfonic acid sultone as the substrate. Both esterase activities could be completely inhibited by adding acetazolamide. In its gross structural characteristics, the C. reinhardii enzyme resembles the CAs from higher plants. However, in its esterase activity and the inhibition by sulfonamides it is markedly different from plant CAs and bears more resemblance to erythrocyte CAs.

Occurrence and some properties of carbonic anhydrases from legume root nodules

Carbonic anhydrase activity (hydration of CO 2 was found in homogenates of leaves (116–500 units.mg −1 protein) and root nodules (27–255 units.mg −1 protein) from 8 legume genera inoculated in each case with a host specific Rhizobium. No enzyme, or only trace amounts (2–7 units.mg −1 protein), were detected in root extracts, The enzymatic activity was inhibited in all cases by azide and acetazolamide. The sizes of nodule and leaf carbonic anhydrases, estimated by gel filtration of partially purified preparations from Phaseolus vulgaris, were around 45 000 and 205 000 respectively. These enzymes also differed in sensitivity to inhibitors. More than 99% of the activity present in Vicia faba nodules was recovered as a soluble enzyme and only a trace was located in the isolated bacteroids.

Purification and properties of carbonic anhydrase from sheep erythrocytes.

ADVERTISEMENT RETURN TO ISSUEPREVArticleNEXTPurification and properties of carbonic anhydrase from sheep erythrocytesRobert J. Tanis and Richard E. TashianCite this: Biochemistry 1971, 10, 26, 4852–4858Publication Date (Print):December 1, 1971Publication History Published online1 May 2002Published inissue 1 December 1971https://doi.org/10.1021/bi00802a004RIGHTS & PERMISSIONSArticle Views94Altmetric-Citations32LEARN ABOUT THESE METRICSArticle Views are the COUNTER-compliant sum of full text article downloads since November 2008 (both PDF and HTML) across all institutions and individuals. These metrics are regularly updated to reflect usage leading up to the last few days.Citations are the number of other articles citing this article, calculated by Crossref and updated daily. Find more information about Crossref citation counts.The Altmetric Attention Score is a quantitative measure of the attention that a research article has received online. Clicking on the donut icon will load a page at altmetric.com with additional details about the score and the social media presence for the given article. Find more information on the Altmetric Attention Score and how the score is calculated. Share Add toView InAdd Full Text with ReferenceAdd Description ExportRISCitationCitation and abstractCitation and referencesMore Options Share onFacebookTwitterWechatLinked InReddit PDF (2 MB) Get e-Alerts Get e-Alerts

Purification and properties of carbonic anhydrase B from porcine erythrocytes

Carbonic anhydrase B (carbonate hydrolase EC 4.2.1.1) has been purified from porcine erythrocytes to at least 90% homogeneity, as measured by sedimentation velocity and electrophoresis at several pH values. The isoelectric point of the enzyme is at 7.30 and its molecular weight by short-column Yphantis sedimentation equilibrium is 30,375 ± 820. The partial specific volume calculated from the amino acid composition is 0.734 cc/g. The sedimentation constant, S 20, w 0, is 2.86 S. From these data, the frictional ratio, f f 0 , is 1.21. The enzyme contains approximately 1 mole of zinc per mole of protein, but no sulfur-containing amino acids or carbohydrate. The amino terminus is acetylated, and the carboxyl terminal sequence of amino acids has been determined by hydrazinolysis and carboxypeptidase A and B digestion to be lys-alaser-phe-COOH. This sequence is more similar to the corresponding sequence from the human enzyme than to that of the bovine.

The purification and properties of carbonic anhydrases from guinea-pig erythrocytes and mucosae of the gastrointestinal tract.

Procedures for isolating carbonic anhydrase (EC 4.2.1.1) enzymes from the erythrocytes and the mucosae of the gastrointestinal tract of guinea pigs are described. From a haemolysate, haemoglobin was removed by the addition of ammonium sulphate, and also by two other methods, namely by gel filtration or by adsorption on DEAE-Sephadex. The crude enzyme thus obtained was resolved into the different isoenzymes by chromatography with DEAE-cellulose. From particle-free supernatants of homogenates of some gastrointestinal tissues, carbonic anhydrases were purified by ammonium sulphate fractionation, gel filtration, and ion-exchange chromatography with DEAE-cellulose. The major isoenzymes from blood, stomach, proximal colonic mucosa and caecal mucosa were homogeneous during ion-exchange chromatography, acrylamide-gel electrophoresis, and centrifugal examination. From these tissues, carbonic anhydrase was isolated as two major isoenzymes. They resemble the pairs of isoenzymes discovered in the bloods of other species. The carbon dioxide hydratase activity of one isoenzyme (;high activity' carbonic anhydrase) was 40 times that of the other isoenzyme (;low activity' carbonic anhydrase), as measured at a single substrate concentration. Two other minor components of the enzyme are also found in guinea-pig erythrocytes. All of the enzymes isolated had molecular weights of nearly 30000 (sedimentation equilibrium). ;High activity' carbonic anhydrases from blood and gastrointestinal tissues were indistinguishable according to some chemical, physical and kinetic measurements; similarly ;low activity' carbonic anhydrases from those tissues were indistinguishable. ;High activity' carbonic anhydrase was markedly different from the ;low activity' carbonic anhydrase with respect to its amino acid composition, chromatographic behaviour and isoelectric pH value. Marked differences were also found in the tissue concentrations of the major isoenzymes. It is suggested that the characteristic and selective distribution of the different forms of carbonic anhydrase in the guinea-pig tissues is related to the specific and different physiological functions of the enzymes.

Purification and properties of carbonic anhydrase from human erythrocytes

1. 1. In fractionation experiments several forms of human erythrocyte carbonic anhydrase were obtained. Purification methods, involving ion-exchange chromatography and zone electrophoresis, are described for the two forms which account for the greatest part of the enzyme activity in ethanol-chloroform extracts of the erythrocytes. 2. 2. The two purified forms of the enzyme show a high degree of homogeneity in electrophoresis and in the ultracentrifuge. Both have a molecular weight of 34 000, but, in contrast to other cases of microheterogeneity, they differ in their specific activities. The enzyme contains 0.36% S and 0.20% Zn, corresponding to 4 atoms of sulfur and 1 atom of zinc per protein molecule. 3. 3. The possible existence of preparation artefacts is discussed and comparisons of the physical and chemical properties of different forms of the human carbonic anhydrase and bovine enzyme are made.