Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.11.12 (PKG)
 2,515 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The structure of the cyclic GMP-binding domain of the cyclic GMP-gated ion channel from bovine retinal rod photoreceptors has been modeled by analogy to the crystal structure of the homologous cyclic AMP-binding domain of catabolite gene activator protein (CAP). The modeled cyclic GMP-binding domain has a three-residue deletion and a five-residue insertion between beta strands compared to CAP. The major interactions of the ion channel with cyclic GMP are similar to those observed for cyclic AMP bound to CAP and predicted for cGMP bound to the cGMP-dependent protein kinase: Gly 543 and Glu 544 make hydrogen-bond interactions with the ribose 2'-OH, Arg 559 forms an ion pair with the charged phosphate oxygen, and Thr 560 forms hydrogen-bond interactions with an exocyclic phosphate oxygen and with the 2-amino group of cGMP. Three additional potential interactions were predicted from the model structure. Ile 545 O and Ser 546 OH form hydrogen-bond interactions with an exocyclic phosphate oxygen, and Phe 533 may interact with the aromatic ring of cGMP. This model is in agreement with both the analogue binding experiments and the mutational analysis of Thr 560.
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PMID:Molecular model of the cyclic GMP-binding domain of the cyclic GMP-gated ion channel. 131 56

cGMP-dependent protein kinase contains four cGMP-binding sites which are homologous to the four cAMP-binding sites of cAMP-dependent protein kinase. The interaction of the diastereomers of adenosine 3',5'-thionophosphate, (PS)-cAMP[S] and (PR)-cAMP[S], with cGMP-dependent protein kinase has been studied. Autophosphorylation of cGMP-dependent protein kinase is stimulated by cAMP and (PS)-cAMP[S] with apparent KA values of 7 microM and 94 microM, respectively. cAMP-stimulated autophosphorylation is inhibited competitively by (PR)-cAMP[S] with a Ki value of 15 microM. The phosphorylation of the peptide substrate (Leu-Arg-Arg-Ala-Ser-Leu-Gly) is stimulated by cGMP (approx. KA 1 microM) and cAMP (approx. KA 98 microM) but neither by the (PR) nor (PS) stereoisomer of cAMP[S]. (PR)-cAMP[S] and (PS)-cAMP[S] inhibit competitively cAMP-or cGMP-stimulated phosphorylation of the peptide substrate with Ki values of 52 microM and 73 microM, respectively. (PS)-cAMP[S] stimulates the phosphorylation of the peptide substrate by an autophosphorylated enzyme. Binding of [3H]cGMP to cGMP-dependent protein kinase is inhibited by (PS)-cAMP[S] and (PR)-cAMP[S] with IC50 values of 200 microM and 15 microM, respectively. These results show that both diastereomers of cAMP[S] bind to cGMP-dependent protein kinase. (PR)-cAMP[S] has properties of a pure antagonist whereas (PS)-cAMP[S] has properties of a partial agonist. The results provide further evidence that autophosphorylation of the enzyme affects the interaction between the cGMP-binding sites and the catalytic center of the enzyme by facilitating the activation of the phosphotransferase reaction.
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PMID:Diastereomers of adenosine 3',5'-monothionophosphate (cAMP[S]) antagonize the activation of cGMP-dependent protein kinase. 299 Sep 28

The specificities of cAMP-dependent and cGMP-dependent protein kinases were studied using synthetic peptides corresponding to the phosphorylation site in 6-phosphofructo-2-kinase/Fru-2,6-P2ase (Murray, K.J., El-Maghrabi, M.R., Kountz, P.D., Lukas, T.J., Soderling, T.R., and Pilkis, S.J. (1984) J. Biol. Chem. 259, 7673-7681) as substrates. The peptide Val-Leu-Gln-Arg-Arg-Arg-Gly-Ser-Ser-Ile-Pro-Gln was phosphorylated by the catalytic subunit of cAMP-dependent protein kinase on predominantly the first of its 2 seryl residues. The Km (4 microM) and Vmax (14 mumol/min/mg) values were comparable to those for the phosphorylation of this site within native 6-phosphofructo-2-kinase/Fru-2,6-P2ase. An analog peptide containing only two arginines was phosphorylated with poorer kinetic constants than was the parent peptide. These results suggest that the amino acid sequence at its site of phosphorylation is a major determinant that makes 6-phosphofructo-2-kinase/Fru-2,6-P2ase an excellent substrate for cAMP-dependent protein kinase. Although 6-phosphofructo-2-kinase/Fru-2,6-P2ase was not phosphorylated by cGMP-dependent protein kinase, the synthetic peptide corresponding to the cAMP-dependent phosphorylation site was a relatively good substrate (Km = 33 microM, Vmax = 1 mumol/min/mg). Thus, structures other than the primary sequence at the phosphorylation site must be responsible for the inability of cGMP-dependent protein kinase to phosphorylate native 6-phosphofructo-2-kinase/Fru-2,6-P2ase. Peptides containing either a -Ser-Ser- or -Thr-Ser- moiety were all phosphorylated by cGMP-dependent kinase to 1.0 mol of phosphate/mol of peptide, but the phosphate was distributed between the two hydroxyamino acids. Substitution of a proline in place of the glycine between the three arginines and these phosphorylatable amino acids caused the protein kinase selectively to phosphorylate the threonyl or first seryl residue and also enhanced the Vmax values by 4-6-fold. These results are consistent with a role for proline in allowing an adjacent threonyl residue to be readily phosphorylated by cGMP-dependent protein kinase.
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PMID:Synthetic peptides corresponding to the site phosphorylated in 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase as substrates of cyclic nucleotide-dependent protein kinases. 300 75

The regulatory subunit of the type I cAMP-dependent protein kinase (Rt) serves as a substrate for the phosphotransferase reaction catalyzed by cGMP-dependent protein kinase (Km = 2.2 microM). The reaction is stimulated by cGMP when RI . cAMP is the substrate, but not when nucleotide-free RI is used. The cGMP-dependent protein kinase catalyzes the incorporation of 2 mol of phosphate/mol of RI dimer in the presence of cAMP and a self-phosphorylation reaction to the extent of 4 mol of phosphate/mol of enzyme dimer. In the absence of cAMP, RI is a competitive inhibitor of the phosphorylation of histone H2B (Ki = 0.25 microM) and of the synthetic peptide substrate Leu-Arg-Arg-Ala-Ser-Leu-Gly (Ki = 0.15 microM) by the cGMP-dependent enzyme. Nucleotide-free RI also inhibits the intramolecular self-phosphorylation of cGMP-dependent protein kinase. The inhibition of the phosphorylation reactions are reversed by cAMP. The catalytic subunit of cAMP-dependent protein kinase does not catalyze the phosphorylation of RIand does not significantly alter the ability of RI to serve as a substrate or an inhibitor of cGMP-dependent protein kinase. These observations are consistent with the concept that the cGMP- and cAMP-dependent protein kinases are closely related proteins whose functional domains may interact.
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PMID:Regulatory subunit of the type I cAMP-dependent protein kinase as an inhibitor and substrate of the cGMP-dependent protein kinase. 624 94

The amino acid sequence around the site of the regulatory subunit of type I cAMP-dependent protein kinase (RI) that is phosphorylated by cGMP-dependent protein kinase has been determined. This site was found to be located near the site on RI previously shown to be very sensitive to hydrolysis by trypsin (Potter, R. L., and Taylor, S. S. (1979) J. Biol. Chem. 254, 2413-2418). The primary sequence surrounding the site is as follows: -Lys-Ala-Gly-Ser-Arg-Ala-Asp-Ser-Arg-Glu-Asp-Glu-Ile-Ser-Pro-Pro-Pro-Pro-Asn-Pro-Val-Val-Lys-Gly-Arg-Arg-Arg-Arg-Gly-Ala-Ile-Ser(P)-Ala-Glu-Val-Tyr-Thr-Glu-Glu-Asp-Ala-Ala-Ser-Tyr-Val-Arg-Lys-Val-Ile-Pro-Lys-Asp-Tyr-Lys-Thr-. As described previously (Geahlen, R. L., and Krebs, E. G. (1980) J. Biol. Chem. 255, 1164-1169), this site is specific for cGMP-dependent protein kinase and is not phosphorylated by the catalytic subunit of cAMP-dependent protein kinase.
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PMID:Studies on the site in the regulatory subunit of type I cAMP-dependent protein kinase phosphorylated by cGMP-dependent protein kinase. 626 84

Chromosomal high mobility group (HMG) proteins have been examined as substrates for cGMP-dependent and cAMP-dependent protein kinases. Of the four HMG proteins only HMG 14 contained a major high affinity site which could be phosphorylated by both enzymes, preferentially by cGMP-dependent protein kinase. One mol of 32P was incorporated/mol of HMG 14. Kinetic analysis revealed apparent Km and Vmax of 40.5 microM and 14.7 mumol/min/mg, respectively, for cGMP-dependent protein kinase, and 123 microM and 11.1 mumol/min/mg, respectively, for cAMP-dependent protein kinase. Tryptic maps of 32P-labeled phosphopeptides of HMG 14 demonstrated phosphorylation of the same site by both enzymes. The tryptic fragment containing the major phosphorylation site was identified by amino acid composition and sequence as HMG 14 (residues 4-13): H-Lys-Val-Ser(P)-Ser-Ala-Glu-Gly-Ala-Ala-Lys-OH. HMG 14 and HMG 17 also contained minor sites which could be phosphorylated by cGMP-dependent protein kinase. Tryptic phosphopeptides mapping suggested that the same minor site was phosphorylated on both HMG 14 and 17. On the basis of amino acid composition, the tryptic peptides carrying the minor phosphorylation sites were identified as H-Leu-Ser(P)-Ala-Lys representing residues 23-26 and 27-30 of HMG 14 and HMG 17, respectively.
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PMID:Phosphorylation of high mobility group 14 protein by cyclic nucleotide-dependent protein kinases. 627 43

The substrate specificity of cGMP-dependent protein kinase has been investigated by examining the ability of the enzyme to phosphorylate a series of synthetic peptides that correspond to the amino acid sequence at its site of autophosphorylation. The undecapeptide Ile53-Gly-Pro-Arg-Thr-Thr58-Arg-Ala-Gln-Gly-Ile63 which corresponds to the sequence around threonine-58 in cGMP-dependent protein kinase (Takio, K., Smith, S.B., Walsh, K.A., Krebs, E.G., and Titani, K. (1983) J. Biol. Chem. 258, 5531-5536) was synthesized and tested as a substrate for that enzyme. It was phosphorylated to the extent of 1.0 mol of phosphate/mol of peptide. Analysis of the products of Edman degradation of the phosphopeptide indicated that only threonine-58 was phosphorylated, as is the case for the autophosphorylation reaction in the native enzyme. The peptide was phosphorylated by cGMP-dependent protein kinase with a Km value of 578 +/- 25 microM and a Vmax of 0.069 +/- 0.003 mumol/min/mg of enzyme. This low Vmax value is consistent with the relatively slow rate of the autophosphorylation reaction. An analog peptide that contained serine in place of threonine-58 was also phosphorylated to 1.0 mol of phosphate/mol of peptide. That phosphopeptide contained only phosphoserine. The serine-containing analog peptide had a Km value similar to that of the parent peptide but was phosphorylated with a 70-fold higher Vmax value. Substitution of arginine-56 in the parent peptide by an alanine residue resulted in a peptide that was essentially not a substrate. Substitution of arginine-59, COOH-terminal to the phosphorylatable threonine, yielded a peptide with a Vmax similar to that of the parent peptide but a Km value of almost 22,000 microM. These results indicate that serine is a better phosphate-accepting residue than is threonine and that both arginine residues around the site of autophosphorylation are important specificity determinants for the cGMP-dependent protein kinase.
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PMID:Phosphorylation by cyclic GMP-dependent protein kinase of a synthetic peptide corresponding to the autophosphorylation site in the enzyme. 631 73

Achatin-I (Gly-D-Phe-Ala-Asp), a tetrapeptide having a D-phenylalanine residue and isolated from Achatina ganglia, has been proposed as an excitatory neurotransmitter of Achatina neurones. In the present study, it was demonstrated using Achatina giant neurones that achetin-I, perfused at alow concentration, enhanced an inward current (Iin) caused by 5-hydroxytryptamine (fast component) and an outward current (Iout) caused by FMRFamide (Phe-Met-Arg-Phe-NH2), and that this peptide suppressed an Iin caused by oxytocin, and Iout caused by acetylcholine and APGW-amide (Ala-Pro-Gly-Trp-NH2). These findings indicate that achatin-I acts not only as a neurotransmitter but also as a neuromodulator for these neurones. In the preliminary experiments, it was shown that an Iin caused by achatin-I on an Achatina giant neurone type, PON (periodically oscillating neurone), was suppressed by H-89 (a PKA inhibitor) and W-7 (calmodulin inhibitor), and that an Iin caused by achatin-I on v-RCON (ventral-right cerebral distinct neurone) was suppressed by KT5823 (PKG inhibitor), suggesting that achatin-I acts on PON via the cyclic AMP-PKA system and on v-RCON via the cyclic GMP-PKG system. Moreover, calmodulin would play a role to produce the Iin for achatin-I on PON via the system mentioned.
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PMID:Further study on the effects of achatin-I, an Achatina endogenous neuroexcitatory tetrapeptide having a D-phenylalanine residue, on Achatina neurones. 885 10

SUMMARY Colletotrichum acutatum causes Key lime anthracnose (KLA) and postbloom fruit drop (PFD) of citrus. We utilized restriction enzyme-mediated integration (REMI) mutagenesis to produce six non-pathogenic mutants from a KLA isolate after screening 1064 transformants on detached Key lime leaves. Subsequently, a gene designated KLAP1 (Key Lime Anthracnose Pathogenicity) was identified from one of the mutants and was demonstrated genetically to be required for pathogenicity to Key lime leaves. The predicted polypeptide encoded by KLAP1 contains a cAMP and cGMP-dependent protein kinase phosphorylation site, and two RGD (Arg-Gly-Asp) cell attachment sequences, a bipartite nuclear targeting sequence, a fungal G-protein alpha subunit signature, a putative metal-binding zinc finger (Cys(2)His(2)) and a putative HMG-I/Y ('high mobility group' non-histone chromatin protein encoding genes) DNA-binding domain (A+T hook), suggesting that KLAP1 may function as a transcription activator in C. acutatum. Sequences homologous to KLAP1 were detected in most C. acutatum isolates examined, and similarity was found in several classes of fungi, animals, plants and bacteria, indicating that KLAP1 is a putative, uncharacterized, conserved transcription activator in fungi. Targeted gene disruption of KLAP1 yielded mutants that were blocked in the penetration stage and were completely defective in pathogenicity on Key lime leaves, but remained pathogenic to flower petals. Complementation of a klap1-null mutant with a full-length KLAP1 gene clone restored complete ability to incite lesions on Key lime. The results indicate that KLAP1 is an important pathogenicity factor in C. acutatum.
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PMID:A gene with domains related to transcription regulation is required for pathogenicity in Colletotrichum acutatum causing Key lime anthracnose. 2056 76