Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.11.11 (AMPK)
 12,425 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Impairment in the stimulation of renal production of 1,25-dihydroxyvitamin D[1,25 (OH)2D] by parathyroid hormone (PTH) occurs in diabetes. Renal response to PTH in terms of 25-hydroxyvitamin D-1-hydroxylase (1-OHase) stimulation involves increased cyclic adenosine monophosphate (cAMP) production, increased cAMP-dependent protein kinase activity, and dephosphorylation of renal ferredoxin (renoredoxin). To identify the step where diabetes might impair PTH stimulation of 1-OHase, we studied the effects of PTH on 1,25(OH)2D production, cAMP content, cAMP-dependent protein kinase activity, and the phosphorylation state of renoredoxin by using renal slices from diabetic and nondiabetic rats. PTH and forskolin significantly stimulated 1,25(OH)2D production in renal slices from nondiabetic animals but not from diabetic animals. PTH-stimulated cAMP production and cAMP-dependent protein kinase activity in renal slices were not altered by diabetes. However, diabetes significantly impaired the capacity of PTH to dephosphorylate renoredoxin and to increase the activity of the 1-OHase enzyme complex. These results suggest that the decreased capacity of PTH to stimulate 1-OHase activity in diabetic animals may reflect the decreased capacity of PTH to alter the phosphorylation state of renoredoxin in these animals.
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PMID:Effects of diabetes mellitus on parathyroid hormone-stimulated protein kinase activity, ferredoxin phosphorylation, and renal 1,25-dihydroxyvitamin D production. 182 5

Adrenodoxin is an iron-sulfur protein which functions as a carrier of reducing equivalents in steroid hydroxylation reactions catalyzed by specific cytochromes P-450 in steroidogenic tissues such as adrenal cortex. Purified bovine adrenocortical adrenodoxin was shown to be selectively phosphorylated upon incubation with purified cAMP-dependent protein kinase, whereas other protein kinases were ineffective. The phosphorylation reaction was completed within 45 min at 30 degrees C and resulted in the optimal incorporation of 1 mol phosphate/mol adrenodoxin. Apoadrenodoxin, lacking the iron-sulfur cluster, was also phosphorylated under similar conditions. An apparent Km of 55 microM with a Vmax of 0.3 pmol 32P incorporated min-1 mg adrenodoxin-1 was calculated. Phosphorylation resulted in a striking change in several molecular properties of adrenodoxin, such as electrophoretic behavior and hydroxyapatite affinity, thus providing the possibility of clearly separating phosphorylated from unphosphorylated adrenodoxin. In addition, phosphoadrenodoxin became refractory to mild trypsin degradation, whereas this was not the case with apoadrenodoxin. The phosphorylated site of adrenodoxin was identified as a serine residue; study of peptide products resulting from CNBr and proteolytic cleavages of phosphoadrenodoxin suggested that Ser-88 was the target of the phosphorylation reaction. The influence of phosphorylation upon adrenodoxin activity was examined using cholesterol side-chain cleavage and 11 beta-hydroxylase (11 beta) systems, reconstituted from purified components. Phosphorylation of adrenodoxin resulted in an average twofold decrease in its Km values for the two specific cytochromes P-450 involved. This effect was paralleled by a positive relationship between the degree of adrenodoxin phosphorylation and its ability to support the overall activity of reconstituted side-chain cleavage and 11 beta-hydroxylase systems. Although it remains to be examined whether adrenodoxin is phosphorylated in the intact cell, the present observations suggest that it represents a potential target in the hormonal regulation of the adrenocortical differentiated functions, especially by stimulatory agents acting through a cyclic-AMP-dependent mechanism, such as adrenocorticotropin.
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PMID:Phosphorylation of bovine adrenodoxin. Structural study and enzymatic activity. 282 99

Parathyroid hormone (PTH) stimulates the renal conversion of 25-OH-vitamin D3 to 1,25-(OH)2-vitamin D3 in young animals. There is evidence that PTH acts via cAMP and cAMP-dependent protein kinase, but the identity of the phosphorylated protein(s) is unknown. The present study investigates the possibility that phosphorylation modification of specific components of the renal mitochondrial, cytochrome P-450-linked 25-OH-vitamin D3-1 alpha-hydroxylase is involved in the regulation of 1,25-(OH)2-vitamin D3 production. Mitochondria were isolated from [32P]phosphate-labeled renal cortical slices which had been divided into control and agonist-treated groups. The hydroxylase protein components from the solubilized mitochondria were partially purified using p-chloroamphetamine-Sepharose affinity chromatography and polyacrylamide gel electrophoresis. Phosphorylation was observed only in a protein with an Mr = 12,000 and a pI of 4.2 by autoradiography of the gels. This radiolabeled protein was immunoprecipitated with adrenodoxin antibody. Additionally, the protein in the same Mr region of the polyacrylamide gel reacted with adrenodoxin antibody and co-migrated with bovine adrenodoxin. PTH and forskolin treatment resulted in decreased phosphate incorporation into the protein, whereas A23187 treatment increased the phosphorylation. In parallel experiments, affinity-isolated hydroxylase from control and PTH-treated slices was used to assess in vitro hydroxylase activity using [3H]25-hydroxyvitamin D3 as substrate. The hydroxylase activity derived from PTH-treated tissue was significantly higher than that of control. From these data, it is proposed that renal response to PTH in terms of 25-hydroxyvitamin D3 hydroxylase stimulation involves dephosphorylation of renoredoxin, the ferrodoxin component of this hydroxylase complex.
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PMID:Parathyroid hormone stimulates dephosphorylation of the renoredoxin component of the 25-hydroxyvitamin D3-1 alpha-hydroxylase from rat renal cortex. 378 51

Adrenalectomy causes a depressed glycogenolytic response to catecholamines in myocardium. Total phosphorylase activity (a + b) is 20% lower in isolated, perfused hearts from adrenalectomized (ADX) rats compared with hearts from sham-operated (sham) rats even though the basal activity ratios (-AMP/+AMP) do not differ. In response to epinephrine (50 nM), the sham group has a higher activity ratio than the ADX group (0.23 vs. 0.16); the difference in specific activities of phosphorylase a in the two groups is even greater, 87 versus 49 U/mg protein. The glycogen content of the heart is 30% lower in the ADX group. Adrenalectomy does not alter the accumulation of cAMP and activation of cAMP-dependent protein kinase caused by epinephrine. Although rat heart contains a heat-stable phosphatase inhibitor, the activity of this inhibitor, as judged by phosphorylase phosphatase activity, is not altered by epinephrine stimulation or by adrenalectomy. Epinephrine perfusion increases the activity ratios (pH 6.8:8.2) of phosphorylase kinase equally in sham and ADX hearts; however, the specific activities of phosphorylase kinase (basal and hormone-stimulated) at either pH are lower after adrenalectomy. The sensitivity of phosphorylase kinase activity to stimulation by calcium is the same in the sham and ADX groups. A radioimmunoassay for phosphorylase kinase detects 10% less of this enzyme in hearts from adrenalectomized animals. Specific activities at pH 6.8 and 8.2 based on the quantity of phosphorylase kinase detected by radioimmunoassay suggest a lower phosphorylation state in the ADX group. Decreases in quantities of phosphorylase and phosphorylase kinase and enzyme dissociation due to glycogen depletion could all contribute to a depressed glycogenolytic response in the ADX group.
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PMID:Effects of adrenalectomy on activation of glycogen phosphorylase in rat myocardium. 608 85

Two key steroidogenic mitochondrial cytochromes P-450 (cholesterol side-chain cleavage (scc) and 11 beta-hydroxylation (11 beta)) were purified from bovine adrenal cortex and examined as potential phosphorylatable substrates using purified cAMP-dependent protein kinase subunit (C) and A type (CKA) and G type (CKG) cAMP-independent casein kinases. Of the two cytochromes P-450, only P-450 11 beta was able to incorporate phosphate from ATP in the presence of C (Km = 7.5 microM), whereas CKA and CKG were ineffective. Phosphorylation of P-450 11 beta (maximum incorporation of 1 mole of 32P per mole of cytochrome, only on serine residues) did not modify the enzymatic activity of an 11 beta-hydroxylation system reconstituted in vitro from purified components, when adrenodoxin was in excess in the reaction. However, kinetic studies showed that P-450 11 beta phosphorylation strikingly increases the P-450 11 beta-adrenodoxin affinity in a phosphorylation-dependent manner. This would result in a net increase in 11 beta-hydroxylase activity under in vivo conditions where adrenodoxin availability is limited. Possible significance of these observations in the regulation of differentiated adrenocortical functions remains to be further examined.
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PMID:Phosphorylation of purified mitochondrial cytochromes P-450 (cholesterol desmolase and 11 beta-hydroxylase) from bovine adrenal cortex. 628 91

Before the structure of cAMP-dependent protein kinase had been solved, sequence alignments had already suggested that several highly conserved peptide motifs described as kinase subdomains I through XI might play some functional role in catalysis. Crystal structures of several members of the protein kinase superfamily have suggested that the nearly invariant aspartate residue within subdomain IX contributes to the conformational stability of the catalytic loop by forming hydrogen bonds with backbone amides within subdomain VI. However, substitution of this aspartate with alanine or threonine in some protein kinases have indicated that these interactions are not essential for activity. In contrast, we show here that conversion of this aspartate to arginine abolished the catalytic activity of the Fer protein-tyrosine kinase when expressed either in mammalian cells or in bacteria. Structural modeling predicted that the catalytic loop of the FerD743R mutant was disrupted by van der Waal's repulsion between the side chains of the substituted arginine residue in subdomain IX and histidine-683 in subdomain VI. The FerD743R mutant model predicted a shift in the peptide backbone of the catalytic loop, and an outward rotation of histidine-683 and arginine-684 side chains. However, the position and orientation of the presumptive catalytic base, aspartate-685, was not substantially changed. The proposed model explains how substitutions of some, but not all residues could be tolerated at this nearly invariant aspartate in kinase subdomain IX.
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PMID:Mutation of a highly conserved aspartate residue in subdomain IX abolishes Fer protein-tyrosine kinase activity. 1019 87

We characterized a regulatory element located in the -76 to -62 region of the human ferredoxin gene. This region bound to Sp1-like proteins with low affinity, as shown using electrophoretic mobility shift, competition, antibody binding, and Southwestern experiments. The similarity of the regulatory element to Sp1 extends beyond its DNA-binding domain, as cloned Sp1 functioned equally well when fused to a peptide that bound to an irrelevant site. The function of these Sp1-binding sites is mediated through the cAMP-dependent protein kinase (PKA) signaling pathway, because reporter genes downstream of the Sp1-binding sites were not activated in a PKA-deficient cell line. Transfection of the catalytic subunit of PKA restored activated transcription. Similar Sp1-binding sites identified in the CYP11A1 and CYP21 genes also controlled cAMP-dependent transcription of the reporter gene. Our finding of the function of Sp1-like proteins in steroidogenic gene transcription adds one more role Sp1 plays in controlling physiological events.
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PMID:Sp1-like proteins function in the transcription of human ferredoxin genes. 1075 89