Abstract
Carbonic anhydrase is an enzyme found in many mammalian tissues, but it has not previously been determined whether it is present in the bovine brain. In this work, carbonic anhydrase was purified and characterized according to localizations: outer peripheral, cytosolic, inner peripheral and integral in four steps. Affinity chromatography was used for purification of the enzyme from the four different cell regions. The affinity column was prepared with Sepharose-4B-l-tyrosine-sulfanilamide. Purified enzymes obtained at each step activity were determined by hydratase activity and esterase activity methods. Optimum pH and optimum temperature values were defined for the purified enzymes. The behavior of carbonic anhydrase with specific inhibitors, sulfanilamide, KSCN and NaN3, was investigated. Molecular weights of enzymes were determined by gel filtration, and its purity controlled by SDS-PAGE electrophoresis. In addition, the enzyme’s K M and V max values were determined with the Lineweaver–Burk method. The results obtained are discussed in comparison with other mammalian carbonic anhydrases.
Similar content being viewed by others
References
Arslan O, Nalbantoğlu B, Demir N, Özdemir H, Küfrevioğlu Öİ (1996) A new method for the purification at carbonic anhydrase isozymes by affinity chromatography. Turk J Med Sci 26:163–166
Bradford MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72:248–254. https://doi.org/10.1016/0003-2697(76),90527-3
Coulson RA, Herbert JD (1984) A role for carbonic anhydrase in intermediary metabolism. Ann N Y Acad Sci 429:505–515. https://doi.org/10.1111/j.1749-6632.1984.tb12379.x
Demir N, Küfrevioğlu Öİ, Keha EE, Bakan E (1993) An enzymatic method for zinc determination in serum. BioFactors 4(2):129–132
Demir Y, Demir N, Bakan E, Küfrevioğlu Öİ, Gündoğdu M (1996) Glycation of granulocyte and monocyte membrane proteins in diabetic patients. Turk J Med Sci 26:467–470
Demir Y, Demir N, Nadaroğlu H, Bakan E (2000) Purification and characterization of carbonic anhydrase from bovine erythrocyte plasma membrane. Prep Biochem Biotechnol 30(1):49–59. https://doi.org/10.1080/10826060008544944
Demir N, Demir Y, Coşkun F, Taşgın E (2012) Expression of cytosolic and noncytosolic carbonic anhydrase enzymes from bovine brain membrane. Asian J Chem 24(4):1511–1514
Dodgson SJ (1991) In: Dodgson SJ, Tashian RE, Gros G, Carter ND (eds) The carbonic anhydrases: cellular physiology and molecular genetics. Plenum Press, New York, pp 3–14
Fujikawa AK, Nishimori I, Taguchi T, Onishi S (1999) Human carbonic anhydrase XIV (CA14): cDNA cloning, mRNA expression, and mapping to chromosome 1. Genomics 61:74–81
Guillaume D, Grisor T, Vergniolle-Burette M (1991) Glial contribution to seizure: carbonic anhydrase activity in epileplic mammalian brain. Epilepsia 32(1):10–15. https://doi.org/10.1111/j.1528-1157.1991.tb05603.x
Laemmli UK (1970) Cleauage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227(5259):680–685. https://doi.org/10.1038/227680a0
Maren TH (1967) Carbonic anhydrase, chemistry, physiology and inhibition. Physiol Rev 47(4):595–781
Maren TH (1984) The general physiology of reactions catalyzed by carbonic anhydrase and their inhibition by sulfonamides. Ann N Y Acad Sci 429:568–579. https://doi.org/10.1111/j.1749-6632.1984.tb12389.x
Rickli EE, Ghazanfar SAS, Gibbons BH, Edsall JT (1964) Carbonic anhydrases from human erythrocytes. J Biol Chem 239:1065–1078
Tasgın E, Nadaroglu H, Demir Y, Demir N (2009) Purification and properties of carbonic anhydrase from bone marrow. Asian J Chem 21:5117–5122
Tobin AJ (1970) Carbonic anhydrase from parsley leaves. J Biol Chem 245:2656–2666
Verpoorte JA, Mehta S, Edsall JT (1967) Esterase activities of human carbonic anhydrase. J Biol Chem 242:4221–4229
Whitaker JP (1963) Determination of molecular weight of proteins by gel filtration on sephadex. Anal Chem 35:1950–1953. https://doi.org/10.1021/ac60205a048
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Ergun, F., Demir, N. Identification of Cytosolic and Noncytosolic Carbonic Anhydrases in Brain. Iran J Sci Technol Trans Sci 42, 1813–1819 (2018). https://doi.org/10.1007/s40995-017-0375-x
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s40995-017-0375-x