Resolution of Adenylate Cyclase Sensitive and Insensitive to Ca2+ and Calcium-dependent Regulatory Protein (CDR) by CDR-sepharose Affinity Chromatography

Overview

Journal Proc Natl Acad Sci U S A

Specialty Science

Date 1979 Jan 1

PMID 284333

Citations 23

Authors

K R Westcott

D C La Porte

D R Storm

Affiliations

Soon will be listed here.

Abstract

Partially purified adenylate cyclase [ATP pyrophosphate-lyase (cyclizing), EC 4.6.1.1] from bovine brain cortex was fractionated into two separate forms by calcium-dependent regulatory protein (CDR)-Sepharose affinity chromatography. The major form of the enzyme, comprising approximately 80% of the applied activity, did not bind to the affinity column in the presence of Ca2+ and was insensitive to the CDR. Approximately 20% of adenylate cyclase activity was absorbed to CDR-Sepharose in the presence of Ca2+. This activity was stimulated by Ca2+ and CDR. This study directly demonstrates that brain cortex contains Ca2+-CDR-sensitive and -insensitive forms of adenylate cyclase and indicates that CDR-Sepharose may be a useful tool for purification of adenylate cyclase. The Ca2+ -stimulated adenylate cyclase was purified at least 55-fold with a 13% yield.

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References

Wallace R, Lynch T, Tallant E, Cheung W . An endogenous inhibitor protein of brain adenylate cyclase and cyclic nucleotide phosphodiesterase. Arch Biochem Biophys. 1978; 187(2):328-34. DOI: 10.1016/0003-9861(78)90042-5. View

Johnson G, Bourne H, Coffino P . Complementation analysis of hormone-sensitive adenylate cyclase. Nature. 1978; 272(5655):720-2. DOI: 10.1038/272720a0. View

Ross E, GILMAN A . Reconstitution of catecholamine-sensitive adenylate cyclase activity: interactions of solubilized components with receptor-replete membranes. Proc Natl Acad Sci U S A. 1977; 74(9):3715-9. PMC: 431700. DOI: 10.1073/pnas.74.9.3715. View

Homcy C, Wrenn S, Haber E . Affinity purification of cardiac adenylate cyclase: dependence on prior hydrophobic resolution. Proc Natl Acad Sci U S A. 1978; 75(1):59-63. PMC: 411183. DOI: 10.1073/pnas.75.1.59. View

Neer E . Size and detergent binding of adenylate cyclase from bovine cerebral cortex. J Biol Chem. 1978; 253(5):1498-502. View

Brostrom C, Brostrom M, Wolff D . Calcium-dependent adenylate cyclase from rat cerebral cortex. Reversible activation by sodium fluoride. J Biol Chem. 1977; 252(16):5677-85. View

Limbird L, Lefkowitz R . Resolution of beta-adrenergic receptor binding and adenylate cyclase activity by gel exclusion chromatography. J Biol Chem. 1977; 252(2):799-802. View

Orly J, Schramm M . Coupling of catecholamine receptor from one cell with adenylate cyclase from another cell by cell fusion. Proc Natl Acad Sci U S A. 1976; 73(12):4410-4. PMC: 431477. DOI: 10.1073/pnas.73.12.4410. View

Klee C, KRINKS M . Purification of cyclic 3',5'-nucleotide phosphodiesterase inhibitory protein by affinity chromatography on activator protein coupled to Sepharose. Biochemistry. 1978; 17(1):120-6. DOI: 10.1021/bi00594a017. View

10.

LaPorte D, Storm D . Detection of calcium-dependent regulatory protein binding components using 125I-labeled calcium-dependent regulatory protein. J Biol Chem. 1978; 253(10):3374-7. View

11.

Watterson D, Vanaman T . Affinity chromatography purification of a cyclic nucleotide phosphodiesterase using immobilized modulator protein, a troponin C-like protein from brain. Biochem Biophys Res Commun. 1976; 73(1):40-6. DOI: 10.1016/0006-291x(76)90494-0. View

12.

Ho H, Wirch E, Stevens F, Wang J . Purification of a Ca2+-activatable cyclic nucleotide phosphodiesterase from bovine heart by specific interaction with its Ca2+-dependent modulator protein. J Biol Chem. 1977; 252(1):43-50. View

13.

Peterson G . A simplification of the protein assay method of Lowry et al. which is more generally applicable. Anal Biochem. 1977; 83(2):346-56. DOI: 10.1016/0003-2697(77)90043-4. View

14.

Takai K, Kurashina Y, HAYAISHI O . Adenylate cyclase from Brevibacterium liquefaciens. Methods Enzymol. 1974; 38:160-9. DOI: 10.1016/0076-6879(74)38025-1. View

15.

Salomon Y, Londos C, Rodbell M . A highly sensitive adenylate cyclase assay. Anal Biochem. 1974; 58(2):541-8. DOI: 10.1016/0003-2697(74)90222-x. View

16.

von Hungen K, Roberts S . Catecholamine and Ca 2+ activation of adenylate cyclase systems in synaptosomal fractions from rat cerebral cortex. Nat New Biol. 1973; 242(115):58-60. DOI: 10.1038/newbio242058a0. View

17.

Johnson R, SUTHERLAND E . Detergent-dispersed adenylate cyclase from rat brain. Effects of fluoride, cations, and chelators. J Biol Chem. 1973; 248(14):5114-21. View

18.

BRADHAM L, Holt D, Sims M . The effect of Ca2+ on the adenyl cyclase of calf brain. Biochim Biophys Acta. 1970; 201(2):250-60. DOI: 10.1016/0304-4165(70)90299-0. View

19.

Lin Y, Liu Y, Cheung W . Cyclic 3':5'-nucleotide phosphodiesterase. Purification, characterization, and active form of the protein activator from bovine brain. J Biol Chem. 1974; 249(15):4943-54. View

20.

Wolff D, Brostrom C . Calcium-binding phosphoprotein from pig brain: identification as a calcium-dependent regulator of brain cyclic nucleotide phosphodiesterase. Arch Biochem Biophys. 1974; 163(1):349-58. DOI: 10.1016/0003-9861(74)90486-x. View