Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:4.6.1.1 (adenylate cyclase)
 19,190 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A technique is described for labeling bovine parathyroid hormone (bPTH) with tritium by [3Hmethyl exchange. The methionine residues were first methylated with [3H]methyl iodide at pH 4, and the reaction products were separated by cation exchange chromatography. The major peak consisted to hormone in which both methionines were converted to [3H]methyl methionine sulfonium iodide (3H-methylated bPTH). This product was then demethylated with 2-mercaptoethanol (6 M) at pH 8.6 to regenerate the hormone in an unmodified but tritiated form ([3H]bPTH), with a specific activity of 1.7 Ci/mmol. High pressure liquid chromatographic analysis showed that 96% of the radioactivity was incorporated into the methionine residues. There was no evidence of any alteration in the primary structure, as [3H]bPTH was found to run in the same position as unlabeled bPTH on cation exchange chromatography and disc gel electrophoresis and to have an identical absorption spectrum in the 240- to 330-nm range. Moreover, [3H]bPTH had full biological activity, as measured by an in vitro bioassay based on activation of rat renal cortical adenylate cyclase, although 3H-methylated bPTH was almost completely inactive. Similarly, while 3H-methylated bPTH had reduced potency in a RIA specific for antigenic sites in the 1--34 region of the sequence, [3H]bPTH was found to have full activity. The preparation of labeled bPTH was repeated using [14C]methyl iodide, with similar results, although [14C]bPTH was found to have somewhat reduced immunological and biological activities. While [3H]bPTH had a lower specific activity than can be obtained by various other techniques for incorporating tritium or 125I into peptides, biosynthetic labeling is at present the only alternative method for preparing biologically active, labeled bPTH without altering the primary structure. By comparison with this technique, the present method gave a product of a much higher specific activity which was labeled specifically in the biologically essential amino-terminal region. The same simple chemical procedures are clearly of wide potential application to the preparation of other labeled peptides.
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PMID:Studies of the biological and immunological properties of parathyroid hormone, labeled selectively on the methionine residues by [3H]methyl exchange to high specific activity. 47 96

Some of the effects of native bovine parathyroid hormone and of the synthetic aminoterminal 1-34 fragment on the adenylate cyclase activity of human fat cell ghosts were studied. Saturating concentrations of both hormone preparations caused a significant increase of enzyme activity by about 200-300%. Guanosine 5'-triphosphate (0.1 mM) inhibited basal enzyme activity but had no substantial effect on parathyroid hormone-stimulated enzyme activity. The guanosine 5'-triphosphate analogue, 5'-guanylyl-imidodiphosphate, produced about a threefold enhancement of basal and parathyroid hormone-stimulated enzyme activities under standard conditions (5 mM Mg+2, 1mM ATP, pH 8.0, 30 degrees C). Activation by parathyroid hormone was not influenced by beta-adrenergic blockade in contrast to stimulation by epinephrine. The sensitivity of the enzyme system to the native and the synthetic parathyroid hormone was, however, abolished after pretreatment of the fat cells with trypsin (1 mg/ml). The stimulatory effects of epinephrine and NaF were not affected by pretreatment with trypsin. The results suggest that human fat cells, like rat adipocytes, contain a multireceptor-coupled adenylate cyclase.
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PMID:Adenylate cyclase of human fat cell ghosts. Stimulation of enzyme activity by parathyroid hormone. 55 1

Three analogues of bovine parathyroid hormone (bPTH), [Tyr-34]bPTH-(1--34) amide, [Nle-8,Nle-18,Tyr-34]bPTH-(1--34)amide, and [Nle-8,Nle-18, o-NPS-Trp-23,Tyr-34]bPTH-(1--34)amide were synthesized by the solid-phase method. The synthetic peptides were found to be homogeneous in multiple analytical systems. These analogues represent the NH2-terminal one-third of the hormone, previously shown to contain all the structural requirements necessary for biological activity, but containing several structural modifications associated with enhancement and stabilization of biological activity. When tested in the in vitro renal adenylylcyclase assay, in which unsubstituted bPTH-(1--34) has a potency of 5400 MRC Units/mg, [Tyr-34]bPTH-(1--34)amide had a potency of 16,1000 MRC Units/mg, [Nle-8,Nle-18,Tyr-34]bPTH-(1--34)amide was 10,100 MRC Units/mg, and [Nle-8,Nle-18,o-NPS-Trp-23,Tyr-34]bPTH-(1--34)amide was 10,600 MRC Units/mg. The diverse structural features incorporated in these hormone analogues, resulting in a several-fold increase in biological activity in vitro, demonstrate additive effects on biological activity and should prove valuable in certain biological applications, as well as in the design of other parathyroid hormone analogues of enhanced potency.
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PMID:Design and synthesis of parathyroid hormone analogues of enhanced biological activity. 56 Sep 53

The effect of prostaglandins and various agents on cyclic adenosine 3',5'-monophosphate generation was studied in isolated rat glomeruli. Specific activity of adenylate cyclase in the glomeruli showed a 12-fold increase over the crude homogenate and a fivefold increase over a tubular preparation. Prostacyclin (PGI2) preferentially stimulated adenylate cyclase of isolated glomeruli at a concentration as low as 10(-9) M. Prostaglandins (PGE1, PGE2, and PGA2) and parathyroid hormone (1-34 synthetic PTH fragment) increased adenylate cyclase in glomeruli with maximal stimulation at 2 X 10(-5) M and 2-4 microgram/ml, respectively. No inhibition of PGE2 on PTH stimulation was observed. Isoproterenol (2 X 10(-4) M) caused a small stimulation, while PGF2alpha, arginine vasopressin, and angiotensin II were ineffective. The presence of guanosine triphosphate in the adenylate cyclase assay enhanced basal and PGE2- and PTH-stimulated activity, but had no effect on NaF stimulation. These findings show an effect of prostaglandins and PTH on the glomerular cAMP system and raise the possibility of a physiological action of these agents on the glomerulus.
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PMID:Stimulation of adenylate cyclase in isolated rat glomeruli by prostaglandins. 72 63

The effects of calcium ion on the adenylate cyclase system was studied in isolated, renal basal-lateral plasma membranes of the rat. Bovine parathyroid hormone (bPTH) and a guanyl triphosphate analogue, Gpp(NH)p were used to stimulate cyclase activity. Under conditions of maximal stimulation, calcium ions inhibited cyclic adenosine monophosphate (cAMP) formation, the formation rate falling exponentially with the calcium concentration. Fifty percent inhibition of either bPTH- or Gpp(NH)p-stimulated activity was given by approximately 50 microns Ca++. Also the Hill coefficient for the inhibition was close to unity in both cases. The concentration of bPTH giving half-maximal stimulation of cAMP formation (1.8 x 10(-8) M) was unchanged by the presence of calcium. These data suggest that calcium acts at some point other than the initial hormone-receptor interaction, presumably decreasing the catalytic efficiency of the enzymic moiety of the membrane complex.
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PMID:Effects of PTH and Ca2+ on renal adenyl cyclase. 74 83

Several aspects of the activation of adenylate cyclase by guanosine 5'-triphosphate (GTP), 5'-guanylylimidodiphosphate (Gpp(NH)p) and bovine parathyroid hormone (bPTH) have been studied in chick kidney plasma membrane preparations. GTP (10(-4) mol/l), Gpp(NH)p (10(-4) mol/l) and bPTH (10 i.u./ml) activated adenylate cyclase without any significant time lag. However a 2 min delay was observed before the activity of the enzyme increased after the addition of bPTH (-6 leads to +34) to incubations. The early (0-3 min) effects of GTP and Gpp(NH)p upon chick kidney adenylate cyclase activity were antagonized by the addition of the alternative guanyl nucleotide. After 5 min of incubation with kidney plasma membranes, Gpp(NH)p induced a stable state of activation of adenylate cyclase which was not reversible by subsequent addition of GTP. GTP did not induce an irreversible state of enzyme activation. In pre-incubation studies, GTP did not produce a persistent enzyme activation and did not modify the effect of Gpp(NH)p added subsequently at the incubation stage. Gpp(NH)p produced a stable state of activation of adenylate cyclase which was not inhibited by addition of GTP at the incubation stage. Bovine PTH (2-34) inhibited the effect of bPTH upon adenylate cyclase activity when the native hormone (10 i.u./ml) had been incubated with plasma membranes for up to 8 min before addition of the analogue (5 mug/ml). Incubation of plasma membranes with bPTH (2-34) for as little as 10 s prevented activation of adenylate cyclase by subsequent addition of bPTH. This pattern was confirmed in pre-incubation studies. After pre-incubation of kidney membranes with bPTH and bPTH (2-34), followed by washing, an acid extract of the membranes contained immunoreactive bPTH. Gpp(NH)p produced a greater increase in adenylate cyclase activity in membranes pre-incubated with bPTH or bPTH (2-34) than in membranes pre-incubated with buffer alone, suggesting that the hormone and analogue facilitated the interaction of Gpp(NH)p with adenylate cyclase.
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PMID:States of activation of chick kidney adenylate cyclase induced by parathyroid hormone and guanyl nucleotides. 83 41

Renal phosphorus handling was evaluated in 12 lithium carbonate-treated psychiatric patients. Serum phosphorus was normal and serum lithium values were within the therapeutic range in all subjects. Serum calcium concentrations measured in 6 of the patients were found to be within the normal range; in the same patients serum parathyroid hormone levels were normal in 4 and slightly elevated in 2. Phosphorus clearance (14 +/- 3 [se] ml/min) and tubular reabsorption of phosphorus (88 +/- 2%) during oral sodium bicarbonate loading were not significantly different from those in 10 healthy control subjects. In a subgroup of 5 patients and 5 control subjects, phosphorus excretion did not increase after bicarbonate loading. In these subjects, phosphorus excretion rates after bicarbonate loading were not different. Although experimental studies suggest that lithium inhibits renal cortical adenylate cyclase stimulation by parathyroid hormone, our data did not indicate any striking effect of long-term lithium administration on serum calcium and serum phosphorus or on renal phosphorus handling.
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PMID:Effect of long-term lithium administration on renal phosphorus handling. 84 75

Rabbit distal convoluted tubules (DCT) microdissected from collagenase-treated kidneys were observed to contain up to four portions of a different appearance under stereomicroscopic examination: (1) a DCTa portion (generally very short), located right after the macula densa (MD) and resembling the portion of the limb (CAL) located before the MD; (2) a constant, "bright" portion, DCTb; (3) a constant, "granular" DCTg portion which, in most DCT, is connected to a portion of the collecting tubule of a similar "granular" appearance (CCTg); (4) many DCT having contacts with the kidney capsule in the superficial cortex were observed to contain an additional portion of a "light" appearance, DCTl, resembling the portion of the collecting tubule (CCTl) to which these superficial DCT are always branched. The hormone-dependent adenylate cyclase (AC) contained in these different portions was investigated by sectioning microdissected distal structures into successive samples according to the above-mentioned criteria, and by measuring with the help of a previously described micromethod, the enzyme activity contained in each single sample under one of the following conditions: control, parathyroid hormone. (PTH l U/ml), vasopressin, (AVP 10(-6)M), isoproterenol (10(-6)M), fluoride (5 X 10(-3)M). Highly significant and reproducible AC stimulations by these hormones were obtained for the following portions, respectively: DCTa, DCTg and CCTg with PTH; DCTl and CCTl with AVP; DCTg, CCTg and CCTl with isoproterenol. From these data, it is concluded that (a) the distal convoluted tubule can no longer be regarded as a single well-defined functional structure; (b) DCTa is actually a short CAL portion extending beyond MD, (c) DCTg and CCTg are two portions of a same functional segment; (d) similarly, DCTl belongs to the functional segment mainly constituted by CCTl; and, finally, (e) DCTb is the only functional segment which is entirely located in the distal convoluted tubule, i.e., included between the macula densa and the first branching with another tubule.
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PMID:Functional segmentation of the rabbit distal tubule by microdetermination of hormone-dependent adenylate cyclase activity. 94 Feb 69

The relative biologic activities of native human parathyroid hormone, hPTH (1-84), native bovine parathyroid hormone, bPTH (1-84), and their respective synthetic, NH2-terminal, biologically-active (1-34) fragments were compared in vitro using adenylate cyclase preparations from human, chicken, and rat renal cortex. The concentrations of the hormones required for half-maximal stimulation of the enzymes were determined from dose response curves. bPTH (1-84) had greater apparent activity than hPTH (1-84), using rat or chicken renal adenylate cyclase, but, with human renal adenylate cyclase, the apparent activities of the two hormones were equal. Synthetic hPTH (1-34) possessed about 1/10 of the apparent activity of hPTH (1-84) in all three adenylate cyclase systems. However, (GLU22)bPTH (1-34) was about equal inapparent activity to bPTH (1-84) in the three systems. We propose that different rates of hormone degradation at or near renal receptor sites may be responsible for the dependence of the relative biologic activity on the assay system used. In the case of hPTH a peptide chain longer than (1-34) may be required for the full biologic activity of the hormone...
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PMID:Relative biologic activities of human and bovine parathyroid hormones and their synthetic, NH2-terminal (1-34) peptides, as evaluated in vitro with renal cortical adenylate cyclase obtained from three different species. 95 43

1-34 N-terminal fragments of human parathyroid hormone with sequences according to Brewer et al (hPTHB) and to Niall et al (hPTHN) were synthesized and compared for their ability to activate bovine and porcine kidney cortex membrane adenylate cyclase. Results show that these two hormone sequences are able to activate these membranes but at least in this in vitro assay, hPTHN is about 10 times more active than hPTHB on bovine as well as on porcine membranes. These "apparent potencies" with respect to the potency of bPTH 1-34 in bovine and porcine membrane assay systems are respectively 4% and 11% for hPTHB and 39% and 156% for hPTHN. These relative potencies may be interpreted as corresponding to species specificity of the hormone receptor structural relationships.
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PMID:Comparative in vitro biological activity of 1-34 N-terminal synthetic fragments of human parathyroid hormone on bovine and porcine kidney membranes. 96 5


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