Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.1.3.16 (calcineurin)
17,112 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

During the purification of annexin VI from pig lung, we previously reported the isolation of another 67 kDa protein (protein 67E) differing from the former by immunological reactivity, amino acid composition, inability to interact with anionic phospholipids in the presence of Ca2+ and inability to inhibit phospholipase A2 [Fauvel, Vicendo, Roques, Ragab-Thomas, Granier, Vilgrain, Chambaz, Rochat, Chap & Douste-Blazy (1987) FEBS Lett. 221, 397-402]. Attempts to phosphorylate protein 67E by the protein tyrosine kinase of epidermal-growth-factor receptor revealed a dramatic inhibition of receptor autophosphorylation, which was also observed with insulin receptor. This inhibitory effect was found to be supported by a phosphatase active towards p-nitrophenyl phosphate, phosphotyrosine, [32P]phosphotyrosyl histones and [32P]phosphotyrosyl poly(Glu,Tyr), but inactive towards phosphoserine, phosphothreonine and [32P]phosphoseryl histones. Although not purified to complete homogeneity, the enzyme was purified 273-fold over EGTA extracts from pig lung and corresponded to a monomeric protein displaying an apparent molecular mass of 67 kDa. With [32P]phosphotyrosyl poly(Glu,Tyr) as substrate, the purified enzyme displayed Km and Vmax. values of 10 microM and 1.93 mumol/min per mg respectively, which compare reasonably well with other recently described phosphotyrosyl protein phosphatases. From these data and from its sensitivity to various inhibitors, it is concluded that protein fraction 67E contains a novel phosphotyrosyl protein phosphatase, the association of which with annexin extract might offer a clue to the understanding of its possible targeting to membrane substrates.
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PMID:Identification, characterization and purification to near-homogeneity of a novel 67 kDa phosphotyrosyl protein phosphatase associated with pig lung annexin extract. 165 82

The protein phosphatase inhibitor okadaic acid was used to investigate the protein phosphatases involved in the endogenous dephosphorylation of proteins in intact synaptosomes. Despite the fact that the calcium-dependent protein phosphatase (calcineurin) is most concentrated in synaptosomes and accounts for approximately 0.3% of synaptoplasmic protein, the majority of the dephosphorylation activity under both basal and depolarisation conditions is due to protein phosphatase type 1 (PP1) and/or protein phosphatase type 2A (PP2A). Nevertheless our results do suggest that calcineurin is active in synaptosomes and has 2 effects: a rapid, direct dephosphorylation of a limited range of substrates and an indirect activation of PP1 presumably by dephosphorylation of protein phosphatase 1 inhibitor-1.
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PMID:Modulation of synaptosomal protein phosphorylation/dephosphorylation by calcium is antagonised by inhibition of protein phosphatases with okadaic acid. 165 31

The mechanism of G protein-mediated sensitization of the contractile apparatus of smooth muscle to Ca2+ was studied in receptor-coupled alpha-toxin-permeabilized rabbit portal vein smooth muscle. To test the hypothesis that Ca2+ sensitization is due to inhibition of myosin light-chain (MLC) phosphatase activity, we measured the effect of guanosine 5'-[gamma-thio]triphosphate and phenylephrine on the rate of MLC dephosphorylation in muscles preactivated with Ca2+ and incubated in Ca(2+)- and ATP-free solution containing 1-(5-chloronaphthalene-1-sulfonyl)-1H-hexahydro-1,4-diazepine (ML-9) to block MLC kinase activity. Guanosine 5'-[gamma-thio]triphosphate alone (300 microM) or in combination (3 microM) with phenylephrine decreased the rates of relaxation and dephosphorylation of MLC to about half of control values; this inhibition is sufficient to account for maximal G protein-mediated Ca2+ sensitization of MLC phosphorylation. The rate of thiophosphorylation of MLC with adenosine 5'-[gamma-thio]-triphosphate was not affected by guanosine 5'-[gamma-thio]triphosphate. We suggest that inhibition of protein phosphatase(s) by G protein(s) may have important regulatory functions.
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PMID:G protein-mediated inhibition of myosin light-chain phosphatase in vascular smooth muscle. 165 67

Chromatography of turkey gizzard extract on Sephacryl S-300 has been shown to fractionate the various smooth muscle phosphatases. We have previously reported the purification and characterization of three of these enzymes, termed smooth muscle phosphatase (SMP)-I, -II, and -IV. Recently, we have purified SMP-III to near homogeneity. Although all of the smooth muscle phosphatases dephosphorylate the isolated myosin light chains, only SMP-III and -IV are active toward intact myosin and, therefore, are most likely to play a direct role in the muscle contraction-relaxation process. SMP-III has a higher molecular weight (390,000), as determined by gel filtration, than the other smooth muscle phosphatases and migrates as single band with a molecular weight of 40,000 in a sodium dodecyl sulfate-polyacrylamide gel. SMP-III is immunologically distinct from SMP-I and -II. It dephosphorylates heavy meromyosin and the isolated myosin light chains at a rapid rate but has low activity toward phosphorylase alpha. The activity of SMP-III is not affected by Ca2+ but is activated by Mn2+.Mg2+ stimulates the activity toward heavy meromyosin but inhibits the myosin light chain phosphatase activity. Attempts to classify SMP-III according to the scheme proposed by Ingebritsen and Cohen (Ingebritsen T. S., and Cohen, P. (1983) Science 221, 331-338) revealed that it is resistant to the heat stable inhibitor-2, suggesting that it is a Type 2 protein phosphatase. However, SMP-III is inhibited by concentrations of okadaic acid which are characteristic of Type 1 protein phosphatases and it binds to heparin-Sepharose like other Type 1 phosphatases. But most interestingly, SMP-III does not dephosphorylate the alpha- or beta-subunits of phosphorylase kinase, a property not reported for any Ser/Thr protein phosphatase.
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PMID:Turkey gizzard smooth muscle myosin phosphatase-III is a novel protein phosphatase. 165 15

We have cloned and sequenced rat testis cDNAs coding for a calcium binding polypeptide similar to calcineurin beta subunit, the Ca(2+)-binding subunit of the Ca2+/calmodulin stimulated protein phosphatase. Rat testis cDNA library was screened with a monoclonal antibody Va1 raised against bovine brain calcineurin beta subunit. The deduced amino acid sequence is similar to that of human brain calcineurin beta subunit with respect to containing four putative calcium binding sites. However, distinct differences were found: 1) The cloned cDNA had six amino acids polypeptide tail at carboxy-terminal which is absent in human brain calcineurin beta subunit. This amino acids tail makes the carboxy-terminal highly hydrophilic in contrast to the human brain beta subunit which is hydrophobic at carboxy-terminal; 2) eleven amino acids at the N terminal of the cloned cDNA were completely different from the corresponding region of the brain calcineurin beta subunit.
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PMID:Isolation and sequence of rat testis cDNA for a calcium binding polypeptide similar to the regulatory subunit of calcineurin. 165 20

Multiple catalytic subunits of the Ca2+ and calmodulin (CaM)-dependent protein phosphatase (PrP) ("calcineurin" or PrP-2B) are derived from at least two structural genes, type 1 ("calcineurin A alpha") and type 2 ("calcineurin A beta "), each of which can produce alternatively spliced transcripts. To examine the possible linkage of these genes, we analyzed genomic DNA from human/hamster hybrid cell lines using probes of 122 base pairs that were designed to bind selectively to exon 3 of the open reading frame. In this region, the nucleotide sequence of the type 2 murine cDNA that we cloned was greater than 99% identical to the type 2 human cDNA but only 78% identical to the type 1 human cDNA. Hybridization to Southern blots containing DNA from all human chromosomes showed that gene 1 was found on chromosome 4, whereas gene 2 segregated to chromosome 10. These data suggest that expression of the two calcineurin genes is not physically linked.
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PMID:Chromosomal mapping of the human genes for the calmodulin-dependent protein phosphatase (calcineurin) catalytic subunit. 165 8

The interaction of calmodulin antagonists with a phosphoprotein phosphatase, calcineurin, was investigated using para-nitrophenyl phosphate (pNPP) as a substrate. Calmidazolium, a potent calmodulin antagonist, inhibited the Ni(2+)-stimulated calmodulin-independent phosphatase activity to much the same extent as it did the Ca2+/calmodulin-stimulated activity. Other calmodulin antagonists, such as trifluoperazine, thioridazine, and W-7, also inhibited the Ni(2+)-stimulated phosphatase activity. On the other hand, calmidazolium only weakly and partially inhibited the Mn(2+)-stimulated phosphatase activity and the other calmodulin antagonists examined increased the Mn(2+)-stimulated activity, in the absence of calmodulin. With the addition of an equimolar amount, as to the inhibited holoenzyme, of the purified B subunit of calcineurin, the Ni(2+)-stimulated phosphatase activity recovered from 38 to 63% of the control level in the presence of 5 microM calmidazolium. When the amount of additional B subunit was increased, the phosphatase activity recovered to 94% of the control level, thereby implying that calmidazolium inhibits the Ni(2+)-stimulated phosphatase activity by interacting with the B subunit, in the absence of calmodulin. The Mn(2+)-stimulated phosphatase activity also recovered from the inhibition by calmidazolium, but a much larger amount of the B subunit was necessary for the recovery. These results indicate that the Ni(2+)- and Mn(2+)-stimulated activities of calcineurin are differentially affected by calmodulin antagonists and that the B subunit plays a crucial role in the expression of the Ni(2+)-stimulated phosphatase activity.
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PMID:Calmodulin antagonists differentiate between Ni(2+)- and Mn(2+)-stimulated phosphatase activity of calcineurin. 166 11

Thiol titrations of bovine brain calcineurin, phosphatase with Ellman's reagent revealed the presence of 5 exposed sulfhydryl groups on the native protein, and 10 sulfhydryl groups on the denatured protein. Attempts were made to identify the location of the free thiols within the catalytic and regulatory domains of the enzyme. Our data indicates that free sulfhydryl groups are absent from the vicinity of the Mg2+ and calmodulin binding sites as well as from the active site of the enzyme. However, the fact that the number of free thiols decreased in the presence of Ca2+ and Mn2+, to 4 and 2 respectively, possibly indicates that either free thiols are at or near these domains or become inaccessible as a consequence of conformational changes induced by the metal ions. The Ca2+ and Ca2+/Mg2+ stimulated activities of calcineurin were monitored during modification with Ellman's reagent, iodoacetate and iodoacetamide. Upon modification of 1-2 of the free thiols the activity of the enzyme increased 1.3 to 10.5-fold depending on the thiol reagent and the stimulatory metal ions employed. Modification of the remainder of the free thiols resulted in a decrease in activity. These results suggest that 1-2 thiols are essential for the full expression of calcineurin activity.
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PMID:Transient activation of calcineurin during thiol modification. 166 19

The oscillatory current response to acetylcholine (ACh) in Xenopus laevis oocytes, mediated by endogenous muscarinic ACh receptors, is known to be mildly desensitized by repetitive ACh applications. Pretreatment of oocytes with staurosporine (an inhibitor of protein kinases) was found not only to abolish this desensitization but also to positively and progressively potentiate oscillatory ACh responses. This sensitization by staurosporine was suppressed by 12-O-tetradecanoylphorbol 13-acetate (an activator of protein kinase C). In staurosporine-untreated (control) oocytes, intracellularly injected calcineurin (an isozyme of Ca2+/calmodulin-dependent protein phosphatase 2B) or Ca2+ enhanced oscillatory ACh responses, while trifluoperazine (a calmodulin inhibitor) suppressed the ACh responses but did not affect oscillatory responses to intracellularly injected inositol 1,4,5-trisphosphate. These results suggest that, as far as short-term changes in receptor responsiveness are concerned, endogenous muscarinic ACh receptors in Xenopus oocytes are desensitized by phosphorylation by protein kinase C and sensitized by dephosphorylation by Ca2+/calmodulin-dependent protein phosphatase 2B.
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PMID:Oscillatory muscarinic acetylcholine responses of Xenopus oocytes are desensitized by protein kinase C and sensitized by protein phosphatase 2B. 166 57

Voltage-activated Ca2+ channel currents were recorded from cultured rat hippocampal neurones using the whole-cell clamp technique with Ba2+ as a charge carrier. After breaking into the cell the amplitude of low-voltage activated Ca2+ channel current increased to a new steady value within 1 min whereas several minutes were required for a full development of the high-voltage activated current (IHVA). Pretreatment of cells with calmodulin antagonists (trifluoperazine or W-13) or protein phosphatase inhibitor, okadaic acid, fastened the development of IHVA. Trifluoperazine (6-40 microM) also increased IHVA when applied after breaking into the cell in standard external solution. Incubation of cells in the presence of permeable precursor of Ca2+ chelator, BAPTA, was without effect. The effects of all inhibitors studied allow to suggest that IHVA in intact cells is largely masked due to activity of calmodulin-activated protein phosphatase.
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PMID:Calmodulin antagonists and protein phosphatase inhibitor okadaic acid fasten the 'run-up' of high-voltage activated calcium current in rat hippocampal neurones. 166 13


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