Adenosine 3′:5′-cyclic monophosphate- and guanosine 3′:5′-cyclic monophosphate-dependent protein kinases: Possible homologous proteins

Abstract

The properties of purified mammalian adenosine 3':5'-cyclic monophosphate (cAMP) and guanosine 3':5'-cyclic monophosphate (cGMP)dependent protein kinases were compared. Several physical characteristics of the two enzymes were similar, including size, shap, affinity for cyclic nucleotide binding, and Km for ATP. In addition, the amino acid compositions of the two proteins indicated a close composition homology (70-90% Adenosine 3':5'-cyclic monophosphate (cAMP)-and guanosine 3':5'-cyclic monophosphate (cGMP)-dependent protein kinases (cAMP kinase and cGMP kinase, respectively) have been described in several vertebrate and invertebrate tissues (1-8). Both enzymes have been purified to homogeneity and the subunit compositions characterized (9-14). While the distribution of the cAMP kinase is ubiquitous in mammalian tissues, there does not appear to be a similar distribution of the cGMP kinase since only lung, cerebellum, heart, and small intestine have appreciable amounts of specific cGMP binding activity (8). On the other hand, cGMP kinase is more widely distributed in the lower species of animals especially in the arthropods where the enzyme was first described (3, 4). Abundant evidence has been provided that indicates that cyclic nucleotide-dependent protein kinases are the major receptors for cyclic nucleotides in eukaryotic cells.Several reports have appeared comparing the properties of cAMP and cGMP kinases from several sources (6-9, 15). In general, these studies emphasized the differences between the two enzymes. One study by Hashimoto et al. (16), however, disclosed similarities in substrate specificity between the two cyclic nucleotide-dependent protein kinases. Since relatively crude enzyme preparations were used, few details concerning substrate specificity and physical nature of the enzymes could be obtained.In this paper, we present data showing that purified mammalian cAMP and cGMP kinases have several similarities in physical properties and substrate specificity. Furthermore, we propose, on the basis of several lines of indirect evidence, that the two enzymes are homologous proteins.MATERIALS AND METHODS Purification of Enzymes. Homogeneous bovine liver catalytic subunit of cAMP kinase (C subunit) was prepared by the method of Sugden et al. (17); homogeneous bovine lung cGMP kinase was purified as described (13,14). The regulatory subunit of cAMP kinase (R subunit) from bovine heart was prepared by a method modified from that of Dills et al. (12)

Cardiac natriuretic peptides act via p38 MAPK to induce the brown fat thermogenic program in mouse and human adipocytes

“…*P < 0.05, **P < 0.01, ***P < 0.001 versus untreated. Typical cycle threshold values in untreated cells are as follows: UCP1, 27; PGC-1α, 23; cytochrome c, 20; and PRDM16, 29. of receptors, they produce closely related chemical messengers that result in the activation of functionally homologous protein kinases: PKA and PKG (45). We previously showed that p38α MAPK is activated in white and brown adipocytes in response to stimulation of β-ARs and cAMP/PKA (46), and p38α MAPK is necessary for transcription of the genes for Ucp1 and Pgc-1α in mice (38).…”

Expression of the catalytic domain of cyclic GMP‐dependent protein kinase in a baculovirus system

“…As shown in Fig 3.2. Substrate spec@ity of the catalytic domain Previous studies from several laboratories have shown that while G-kinase demonstrates overlapping substrate specificity with several other protein kinases, especially the A-kinase, there appear to be specificity determinants for the G-kinase as well [20,21]. Nevertheless, the molecular basis for specificity is not well-defined and may relate to the recognition of specific sequence motifs in substrates by the catalytic domain, by the targeting of the holoenzyme to subcellular structures containing substrates for protein kinases, or both.…”

“…Ten ~1 of the extract was assayed for G-kinase in 100 ,ul of assay buffer consisting of 50 mM Tris-HCl, pH 7.5, 20 mM magnesium acetate, 0.2 mM [Y-~*P]ATP (200 cpm/ pmol), and either histone F2b (0.1 mg/ml) or 100 PM of peptide substrate selective for G-kinase, RKISASEFDRPL, initially described by Colbran et al [19]. All assays for G-kinase were performed in the presence of 0.9pM protein kinase inhibitor peptide (5)(6)(7)(8)(9)(10)(11)(12)(13)(14)(15)(16)(17)(18)(19)(20)(21)(22)(23)(24). Assays were conducted at 30°C for 5 min and terminated by aliquoting histone or the peptide to Whatman 3MM filter paper or P81 phosphocellulose paper, respectively.…”

Interruption of Homologous Desensitization in Cyclic Guanosine 3′,5′-Monophosphate Signaling Restores Colon Cancer Cytostasis by Bacterial Enterotoxins

“…Tachyphylaxis induced by heat-stable enterotoxins was not altered by inhibition of PDE5 or application of 8-br-cGMP, suggesting that cGMP hydrolysis is not central to this process. Rather, 8-br-cGMP, a potent activator of PKG (44), induced tachyphylaxis to acute cGMP signaling. In addition, removal of extracellular Ca 2+ from the medium did not affect the development of tachyphylaxis to heat-stable enterotoxins, indicating that Ca 2+ entering through CNG channels and the associated downstream intracellular effectors did not mediate acute desensitization.…”