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Factors affecting the binding of [3H]adenosine 3':5'-cyclic monophosphate to protein kinase from bovine adrenal cortex. 1977

S O Doskeland, and P M Ueland, and H J Haga

Inorganic salts, several proteins and traces of protein precipitants were tested to find out by what mechanisms they modulate the binding of cyclic [3H]AMP to protein kinase (ATP-protein phosphotransferase; EC 2.7.1.37). The separation of free and bound cyclic AMP by (NH4)2SO4 precipitation was unaffected by the above agents and was more reliable than the Millipore filtration technique. Several binding sites for cyclic AMP were revealed in adrenal-cortex extract. When this extract was used as binding reagent in an assay for cyclic AMP, the standard curve was distorted in the presence of KCl because the salt affected the different binding sites to a varying extent. At high ionic strenth the protein kinase isoenzyme I dissociated and showed an extraordinarily high affinity for cyclic AMP. Trichloroacetate and perchlorate at very low concentrations were able to dissociate the protein kinase and modulate its binding characteristics as well. A progressive decrease in the cyclic AMP-binding capacity occurred on prolonged incubations. The binding protein was protected against inactivation by 2-mercaptoethanol, EDTA and several proteins. It was more resistant to denaturation when complexed to cyclic AMP. The enhancement of cyclic AMP binding by bovine serum albumin was investigated in some detail and appeared to be a pure stabilizing effect. It is proposed that the competitive-binding assays for cyclic AMP based on protein kinase be conducted at high ionic strength and in the presence of stabilizers (protein, EDTA, 2-mercaptoethanol). The interference from agents that may dissociate the protein kinase or influence its stability will thus be decreased.

UI MeSH Term Description Entries
D009994 Osmolar Concentration The concentration of osmotically active particles in solution expressed in terms of osmoles of solute per liter of solution. Osmolality is expressed in terms of osmoles of solute per kilogram of solvent. Ionic Strength,Osmolality,Osmolarity,Concentration, Osmolar,Concentrations, Osmolar,Ionic Strengths,Osmolalities,Osmolar Concentrations,Osmolarities,Strength, Ionic,Strengths, Ionic
D011232 Chemical Precipitation The formation of a solid in a solution as a result of a chemical reaction or the aggregation of soluble substances into complexes large enough to fall out of solution. Precipitation, Chemical
D011485 Protein Binding The process in which substances, either endogenous or exogenous, bind to proteins, peptides, enzymes, protein precursors, or allied compounds. Specific protein-binding measures are often used as assays in diagnostic assessments. Plasma Protein Binding Capacity,Binding, Protein
D011494 Protein Kinases A family of enzymes that catalyze the conversion of ATP and a protein to ADP and a phosphoprotein. Protein Kinase,Kinase, Protein,Kinases, Protein
D002417 Cattle Domesticated bovine animals of the genus Bos, usually kept on a farm or ranch and used for the production of meat or dairy products or for heavy labor. Beef Cow,Bos grunniens,Bos indicus,Bos indicus Cattle,Bos taurus,Cow,Cow, Domestic,Dairy Cow,Holstein Cow,Indicine Cattle,Taurine Cattle,Taurus Cattle,Yak,Zebu,Beef Cows,Bos indicus Cattles,Cattle, Bos indicus,Cattle, Indicine,Cattle, Taurine,Cattle, Taurus,Cattles, Bos indicus,Cattles, Indicine,Cattles, Taurine,Cattles, Taurus,Cow, Beef,Cow, Dairy,Cow, Holstein,Cows,Dairy Cows,Domestic Cow,Domestic Cows,Indicine Cattles,Taurine Cattles,Taurus Cattles,Yaks,Zebus
D000242 Cyclic AMP An adenine nucleotide containing one phosphate group which is esterified to both the 3'- and 5'-positions of the sugar moiety. It is a second messenger and a key intracellular regulator, functioning as a mediator of activity for a number of hormones, including epinephrine, glucagon, and ACTH. Adenosine Cyclic 3',5'-Monophosphate,Adenosine Cyclic 3,5 Monophosphate,Adenosine Cyclic Monophosphate,Adenosine Cyclic-3',5'-Monophosphate,Cyclic AMP, (R)-Isomer,Cyclic AMP, Disodium Salt,Cyclic AMP, Monoammonium Salt,Cyclic AMP, Monopotassium Salt,Cyclic AMP, Monosodium Salt,Cyclic AMP, Sodium Salt,3',5'-Monophosphate, Adenosine Cyclic,AMP, Cyclic,Adenosine Cyclic 3',5' Monophosphate,Cyclic 3',5'-Monophosphate, Adenosine,Cyclic Monophosphate, Adenosine,Cyclic-3',5'-Monophosphate, Adenosine,Monophosphate, Adenosine Cyclic
D000302 Adrenal Cortex The outer layer of the adrenal gland. It is derived from MESODERM and comprised of three zones (outer ZONA GLOMERULOSA, middle ZONA FASCICULATA, and inner ZONA RETICULARIS) with each producing various steroids preferentially, such as ALDOSTERONE; HYDROCORTISONE; DEHYDROEPIANDROSTERONE; and ANDROSTENEDIONE. Adrenal cortex function is regulated by pituitary ADRENOCORTICOTROPIN. Cortex, Adrenal
D000311 Adrenal Glands A pair of glands located at the cranial pole of each of the two KIDNEYS. Each adrenal gland is composed of two distinct endocrine tissues with separate embryonic origins, the ADRENAL CORTEX producing STEROIDS and the ADRENAL MEDULLA producing NEUROTRANSMITTERS. Adrenal Gland,Gland, Adrenal,Glands, Adrenal
D000418 Albumins Water-soluble proteins found in egg whites, blood, lymph, and other tissues and fluids. They coagulate upon heating. Albumin
D000818 Animals Unicellular or multicellular, heterotrophic organisms, that have sensation and the power of voluntary movement. Under the older five kingdom paradigm, Animalia was one of the kingdoms. Under the modern three domain model, Animalia represents one of the many groups in the domain EUKARYOTA. Animal,Metazoa,Animalia

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