Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.1.3.16 (calcineurin)
 17,112 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The effects of ATP and divalent cations on a divalent cation-independent phosphorylase phosphatase of Mr = 35,000 (phosphatase S) purified from canine cardiac muscle have been studied. The enzyme can be rapidly inactivated by ATP or other nucleoside di- and triphosphates and PPi, but not by AMP, adenosine, adenine, Pi, EDTA, ethylene glycol bis(beta-aminoethyl ether)N,N' -tetraacetic acid, 1,10-phenanthroline, or 8-hydroxyquinoline. After removing the inactivating agent, such as ATP or PPi, by gel filtraiton followed by exhaustive dialysis, the inactivated enzyme (apophosphatase S) can be reactivated by preincubating with Mn2+ or Co2+, but not with Mg2+, Ca2+, Ni2+, Zn2+, Fe2+, Cu2+, Ba2+, Hg2+, Pb2+, or Cd2+. The Mn2+ -reactivated enzyme, which is less active than the Co2+ -reactivated enzyme, can be again inactivated by preincubating with ATP. The present findings indicate that phosphatase S contains a tightly bound divalent cation, probably Mn2+, in the active site. ATP and PPi, due to their structural similarity to the phosphoprotein substrate and their ability to chelate metal ions, can readily enter the active site to remove the divalent cation(s) essential for the catalytic function. The present findings also indicate that phosphatase S, a common catalytic subunit of several larger molecular forms of nospecific phosphoprotein phosphatase in cardiac muscle, can exist in two interconvertible forms, a metallized form (active) and a demetallized form (inactive). ATP and metal ions may regulate this class of isozymes by mediating the interconversions.
 ...
PMID:The role of ATP and divalent cations in the regulation of a cardiac phosphorylase phosphatase (phosphoprotein phosphatase) of Mr = 35,000. 21 Nov 35

The effects of divalent metals, metal chelators (EDTA, EGTA) and sodium dodecyl sulfate were investigated on the phosphatase activity of isolated bovine brain calcineurin assayed in the absence (called intrinsic) and presence of calmodulin. Intrinsic phosphatase was increased by Mn2+, was unaffected by Mg2+, Ca2+, and Ba2+, and was markedly inhibited by Ni2+, Fe2+, Zn2+ and Cu2+. When assayed in the presence of calmodulin, many divalent metals (Ni2+, Zn2+, Pb2+, Cd2+), besides Mn2+, increased modestly the phosphatase activity at low concentrations (10-100 microM) and inhibited it markedly at high concentrations. Ca2(+)-calmodulin stimulated phosphatase activity was antagonized by Ni2+, Zn2+, Fe2+, Cu2+, Pb2+, at low concentrations (50 microM), and by Ba2+, Cd2+ at slightly higher concentrations (greater than 100 microM); Mn2+ and Co2+ (50 microM to 1 mM) in fact augmented it. EDTA and EGTA in a concentration and time dependent fashion inhibited the intrinsic phosphatase activity, particularly that of trypsinized calcineurin. SDS in low concentrations (0.005%) augmented the phosphatase activity and inhibited it at high concentrations. Mn2+ (+/- calmodulin) and Ca2+ only with calmodulin present increased the phosphatase activity assayed with low concentrations of SDS. The EDTA dependent inhibition of intrinsic phosphatase was almost abolished in assays containing SDS. Prior exposure of calcineurin to Mn2+ led to a high activity conformation state of calcineurin that was 'long-lived' or 'pseudo-irreversible'. Such Mn2(+)-activated state of calcineurin exhibited no discernible change in the affinity towards myelin basic protein or its inhibition by trifluoperazine. At alkaline pH, Mg2+ supported the intrinsic phosphatase activity, although to a lesser degree than Mn2+. The latter cation, compared to Mg2+ and Ni2+, was also a more powerful stimulator of the calcineurin phosphatase assayed with histone (III-S) and myosin light chain as substrates.
 ...
PMID:Divalent cation effects on calcineurin phosphatase: differential involvement of hydrophobic and metal binding domains in the regulation of the enzyme activity. 170 Oct 13

Ultraviolet (280-nm) irradiation of bovine brain calmodulin results in calcium-dependent changes in its fluorescence emission spectrum. These consist of a decline in the intrinsic tyrosine fluorescence of the protein and the appearance of a new emission maximum at 400 nm. Chromatography of irradiated calmodulin, using Ultrogel AcA 54 and phenyl-agarose columns, yields several distinctive fractions. One of these, representing 2.8% of the total recovered protein and 53% of the total fluorescence emission at 400 nm, was selected for detailed characterization. Analyses performed on acid hydrolysates reveal the presence of dityrosine, a derivative of tyrosine known for its fluorescence near 400 nm, at the level of 0.59-0.89 mol per 16,700 g of protein. Sodium dodecyl sulfate gel electrophoresis experiments demonstrate two components of apparent molecular weights 14,000 (80%) and 16,000 (20%). Observations on the effects of UV irradiation on the thrombic fragments of calmodulin and on related calcium binding proteins (rabbit skeletal muscle troponin C, bovine cardiac troponin C, and parvalbumin) support the interpretation that dityrosine formation in calmodulin results from the intramolecular cross-linking of Tyr-99 and Tyr-138. The dityrosine-containing photoproduct of calmodulin is unable to stimulate the p-nitrophenyl phosphatase activity of calcineurin under standard assay conditions. Fluorescence titrations show a generally weakened interaction with calcium ion occurring in two stages. The pKa of the derivative is considerably higher than that of free dityrosine and is calcium dependent, decreasing from 7.88 to 7.59 on the addition of 3 mM CaCl2. Smooth muscle myosin light chain kinase binds the derivative about 280-fold less effectively than it binds native calmodulin. Of several metal ions tested, only Cd2+ approaches Ca2+ in its ability to promote the appearance of the 400-nm emission band during UV irradiation of calmodulin. Mn2+ and Cu2+ appear to inhibit dityrosine formation. Ascorbic acid, dithiothreitol, and glutathione are also inhibitory.
 ...
PMID:Dityrosine formation in calmodulin. 356 41

Phosphoprotein phosphatases (phosphoprotein phosphohydrolase, EC 3.1.3.16) were partially purified from bovine thyroid with phosphorylated mixed histones, H1 histone and casein as substrates. Utilizing DEAE-cellulose chromatography, (NH4)2SO4 precipitation, gel filtration before and after freeze-thawing in 0.2 M 2-mercaptoethanol and histone-Sepharose chromatography, four fractions of enzyme activity were obtained and were designated as phosphatases I, IIA, IIB, and III. Phosphatases I had an apparent molecular weight of 155,000 and was dependent on Mn2+ for maximal activity. The enzyme had the greatest activity with histone H1 and was greatly stimulated by NaCl with phosphohistones as substrate. Phosphatases IIA and IIB had a molecular weight of about 70,000, were stimulated over 5-fold by Mn2+ and had much higher activities with phosphohistones than with casein in the presence of the cation. Phosphatase III, a possible catalytic subunit of larger molecular weight forms, had an apparent molecular weight of 30,000, was generally independent of Mn2+ and had high activities using all three substrates. Phosphatases I, IIA, and III were inhibited in a dose-dependent manner by sodium pyrophosphate (PPi), ATP, potassium phosphate (Pi) and sodium fluoride (NaF) when they were added directly to the reaction mixture with phosphorylated mixed histones as substrate. PPi was the most potent inhibitor and phosphatase III was the most sensitive to inhibition. PPi, ATP and NaF probably inactivated phosphatase III activity by removing an essential metal ion. After extensive dialysis to remove these inhibitors, the inactivated enzyme could be fully activated by Mn2+, but not by Mg2+, Ba2+, Cu2+, Cd2+, Ca2+, Zn2+ and Fe2+. Whereas the enzyme pretreated with Pi retained about 80% activity after dialysis, its activity was not further stimulated by Mn2+. The inactivated (demetallized) enzyme was less reactivated by Mn2+ in the presence of mM concentration of Pi. Moreover, the Mn2+-reactivated enzyme was again inactivated by Pi, NaF and ATP. Among them Pi was the most potent inactivator. These results suggest that Pi may have another inhibitory effect on metal ion binding besides on substrate binding and also that phosphatase III might be a metalloenzyme. In bovine thyroid, there are at least two major phosphoprotein phosphatases which may have different properties. Metal ion stimulation of phosphatase I and IIA activities may be through an interaction with the substrate or with a metal ion binding site on the regulatory subunit. The lowest molecular weight enzyme (phosphatase III) probably does not exist naturally in the cell.
 ...
PMID:Discrimination of multiple forms of phosphoprotein phosphatase in bovine thyroid. 629 68

The catalytic subunit of the major protein phosphatase associated with bovine cardiac myofibrils was purified to homogeneity. Sodium dodecyl sulfate polyacrylamide gel electrophoresis of the enzyme revealed only one band with an apparent molecular weight of 37,000. On gel filtration chromatography, the phosphatase activity and the protein co-eluted as a single peak with an apparent molecular weight of 37,000. The purified enzyme was identified as the catalytic subunit of protein phosphatase 1, as determined by sensitivity to inhibitor 1, inhibitor 2, okadaic acid and by specific immunostaining. Evidence obtained with specific antipeptide antibodies demonstrated that this myofibril protein phosphatase was predominantly the alpha isoform of protein phosphatase 1. The purified catalytic subunit was completely inactive. It was activated by pretreatment with Co2+/trypsin in the presence of high ionic strength. Treatment with trypsin alone did not activate the latent enzyme. The enzyme was also activated by Co2+ or Mn2+ alone but not by Ca2+, Mg2+, Ni2+, Cu2+ or Zn2+. Activation of the enzyme was not reversed by removal of Co2+, but Mn(2+)-activated phosphatase activity was partially reversed when Mn2+ was removed. The catalytic subunit could form a 1:1 complex with inhibitor 2 in vitro. The resulting holoenzyme was also activated by pretreatment with Co2+. Since phosphatase 1 alpha is the major phosphatase associated with cardiac myofibril, it is suggested that it is responsible for the dephosphorylation of myosin and other myofibril phosphoproteins.
 ...
PMID:A latent form of protein phosphatase 1 alpha associated with bovine heart myofibrils. 808 38

We have observed that soluble extracts from the extreme acidothermophilic archaebacterium Sulfolobus solfataricus contained protein phosphatase activity that was greatly stimulated by the divalent metal ions Mn2+, Mg2+, Ni2+, or Co2+. This activity apparently arose from a single enzyme since (a) stimulation by these divalent metal ions was not additive and (b) protein phosphatase activity eluted as a single peak from both a DE52 ion-exchange column and a Sephadex G-100 gel filtration column. Its apparent molecular mass was approximately 28,000 daltons. The enzyme dephosphorylated a variety of phosphoserine-containing substrates including casein, histone H2a, phosphorylase kinase, or glycogen phosphorylase. The enzyme would not dephosphorylate either histone H1 or a number of phosphotyrosine-containing compounds. It removed only half the phosphate bound to histone H2b, which is phosphorylated at two sites by the cAMP-dependent protein kinase. Protein phosphatase activity was inhibited by EDTA, Cu2+, Zn2+, NaF, inorganic phosphate, or pyrophosphate; but was unaffected by other potential activators and inhibitors such as microcystin, okadaic acid, vanadate, polyamines, or sulfhydryl modifying reagents. This enzyme represents the first protein phosphatase to be identified in any member of the third and oldest phylogenetic kingdom in nature, the archaebacteria.
 ...
PMID:Identification of a serine/threonine-specific protein phosphatase from the archaebacterium Sulfolobus solfataricus. 838 14

The recombinant catalytic subunit of protein phosphatase 1 is produced as an inactive enzyme which can be activated by Mn2+ (Zhang, Z., Bai, G., Deans-Zirattu, S., Browner, M. F., and Lee, E. Y. C. (1992) J. Biol. Chem. 267, 1484-1490). In this report, we have investigated the effects of divalent cations on the activity of recombinant catalytic subunit of protein phosphatase 1. Latent phosphatase 1 can be activated by Co2+ or Mn2+, whereas other metal ions tested including Fe2+, Zn2+, Mg2+, Ca2+, Cu2+, or Ni2+ were not effective or were only weakly effective in activating the enzyme. The Mn(2+)-stimulated activity was susceptible to inactivation by EDTA; however, the Co(2+)-activated phosphatase was stable after dilution and chelation of the Co2+ with excess EDTA. After stable activation of phosphatase 1 using 57Co2+, a stoichiometric amount of 57Co2+ was shown to be tightly bound to phosphatase 1. These findings demonstrate for the first time the generation of a stable metalloenzyme form of phosphatase 1. Fe2+ reversibly deactivated the Co(2+)-stimulated activity, but did not displace the bound Co2+. Interestingly, treatment of the enzyme with a combination of Fe2+ and Zn2+ (but not the individual metal ions) significantly activated phosphatase 1. These results suggest that at least two metal binding sites exist on the enzyme and that protein phosphatase 1 may be an iron/zinc metalloprotein in vivo.
 ...
PMID:Activation of protein phosphatase 1. Formation of a metalloenzyme. 857 23

Analysis of 94 kb of DNA, located between map positions 88 and 182 kb in the 330-kb chlorella virus PBCV-1 genome, revealed 195 open reading frames (ORFs) 65 codons or longer. One hundred and five of the 195 ORFs were considered major ORFs. Twenty-six of the 105 major ORFs resembled genes in the databases including three chitinases, a chitosanase, three serine/threonine protein kinases, two additional protein kinases, a tyrosine protein phosphatase, two ankyrins, an ornithine decarboxylase, a copper/zinc-superoxide dismutase, a proliferating cell nuclear antigen, a DNA polymerase, a fibronectin-binding protein, the yeast Ski2 protein, an adenine DNA methyltransferase and its corresponding DNA site-specific endonuclease, and an amidase. The genes for the 105 major ORFs were evenly distributed along the genome and, except for one noncoding 1788-nucleotide stretch, the genes were close together. Unexpectedly, a 900-bp region in the 1788-bp noncoding sequence resembled a CpG island.
 ...
PMID:Analysis of 94 kb of the chlorella virus PBCV-1 330-kb genome: map positions 88 to 182. 861 77

Saccharomyces cerevisiae Lys7p was proposed to be the enzyme catalyzing the dehydratation of homocitrate to cis-homoaconitate, the second step of the lysine biosynthetic pathway. In this communication we provide evidence that Lys7p is involved in oxidative stress protection. Cells deleted for the LYS7 gene displayed, in addition to lysine auxotrophy, methionine auxotrophy, sensitivity to superoxide generating drugs and light irradiation, and diminution of calcineurin activity. The SOD1 gene encoding the Cu/Zn-superoxide dismutase was expressed in strains lacking Lys7p, and although Sodlp was produced in normal amounts no detectable enzyme activity was found. In contrast, the mitochondrial Mn-superoxide dismutase activity did not seem to be impaired. lys7 cells exhibited a normal uptake of Cu from growth medium. The Cu/Zn-superoxide dismutase activity was restored by addition of Cu (but not by addition of other metallic cations) to the growth medium or to cellular extracts, suggesting a lack of Cu2+ at the active site. These results render it necessary to reconsider the role of the Lys7p. Its involvement in Cu metabolism and oxidative-stress protection, and the possibility of a human equivalent in amyotrophic lateral sclerosis are discussed.
 ...
PMID:The Saccharomyces cerevisiae LYS7 gene is involved in oxidative stress protection. 949 44

Freshly isolated protein phosphatase 2A (PP2A) was highly active as to the dephosphorylation of protein substrates, but lost most of its spontaneous activity on prolonged storage, and was converted to a latent form requiring Mn2+ or Co2+ ions for activity. In this report, we show that the latent form of PP2A can be activated by the Fe2+/ascorbate system. Activation of the phosphatase required both Fe2+ ions and ascorbate, and the level of activation was dependent on the concentrations of both Fe2+ ions and ascorbate. Both the holoenzyme and catalytic subunit of phosphatase 2A could be activated by the Fe2+/ascorbate system, indicating that direct modulation of the catalytic subunit of the phosphatase by the Fe2+/ascorbate system may cause this activation. Several common divalent metal ions, including Ca2+, Mg2+, Cu2+, Zn2+, and Ni2+ ions, cannot cooperate with ascorbate to activate the phosphatase. Dithiothreitol, a SH-containing reducing agent, could replace ascorbate in the Fe2+/ascorbate system to activate the phosphatase, whereas H2O2, a strong oxidizer, significantly diminished the phosphatase activation by the Fe2+/ascorbate system. The results indicate that iron ions stabilized in the +2 state by reducing agents can activate the phosphatase. Overall, the present study provides initial biochemical evidence suggesting that Fe2+ could be a biologically important metal ion cofactor responsible for PP2A activation.
 ...
PMID:Activation of protein phosphatase 2A by the Fe2+/ascorbate system. 964 67


1 2 3 4 Next >>