• Microbiology and Biotechnology Letters
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• v.33 no.3
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• pp.189-193
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• 2005

IgE-dependent histamine-releasing factor(HRF) was initially described as a secretagogue for secretion of histamine from IgE+ basophils from a subset of allergic donors. Previously, we identified that S98 residue of HRF was phosphorylated using anti-HRFpS98 antibody which specifically recognizes the phosphorylated serine residue of HRF and HRFS98A mutant construct. In vitro kinase assay, only wild type HRF was phosphorylated by PKC, and S98A HRF was not affected by PKC. In this study, we attempted to characterize the phosphatase which specifically dephosphorylates HRF by immunoprecipitation and pull-down assay. In RBL-2H3 cells, HRF interacted only with calcineurin (also called as PP2B, calcium/calmodulin-dependent phosphatase) but not with PP1 or PP2A. The results suggest that HRF is most likely dephosphory-lated by calcineurin.

• The Plant Pathology Journal
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• v.28 no.3
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• pp.259-269
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• 2012

Protein phosphatase 2A (PP2A), a family of serine/threonine protein phosphatases, plays an important role in balancing the phosphorylation status of cellular proteins for regulating diverse biological functions in eukaryotic organisms. Despite intensive studies in mammals, limited information on its role is available in filamentous fungi. Here, we investigated the functional roles of genes for a putative B' delta regulatory subunit (FgPP2AR) and a catalytic subunit (FgPP2AC) of PP2A in a filamentous ascomycete, Fusarium graminearum. Molecular characterization of an insertional mutant of this plant pathogenic fungus allowed us to identify the roles of FgPP2AR. Targeted gene replacement and complementation analyses demonstrated that the deletion of FgPP2AR, which was constitutively expressed in all growth stages, caused drastic changes in hyphal growth, conidia morphology/germination, gene expression for mycotoxin production, sexual development and pathogenicity. In particular, overproduction of aberrant cylindrical-shaped conidia is suggestive of arthroconidial induction in the ${\Delta}FgPP2AR$ strain, which has never been described in F. graminearum. In contrast, the ${\Delta}FgPP2AC$ strain was not significantly different from its wild-type progenitor in conidiation, trichothecene gene expression, and pathogenicity; however, it showed reduced hyphal growth and no perithecial formation. The double-deletion ${\Delta}FgPP2AR;{\Delta}FgPP2AC$ strain had more severe defects than single-deletion strains in all examined phenotypes. Taken together, our results indicate that both the putative regulatory and catalytic subunits of PP2A are involved in various cellular processes for fungal development in F. graminearum.

• Bulletin of the Korean Chemical Society
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• v.18 no.4
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• pp.428-432
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• 1997

The dephosphorylation specificity of protein phosphatase 2A (PP2A), calcineurin (PP2B) and protein phosphatase 2C (PP2C) were studied in vitro using myelin basic protein (MBP) as a model substrate which was fully phosphorylated at multiple sites by protein kinase C (PKC) or cyclic AMP-dependent protein kinase (PKA). In order to determine the site specificity of phosphates in myelin basic protein, the protein was digested with trypsin and the radioactive phosphopeptide fragments were isolated by high performance liquid chromatography (HPLC) on reversed-phase column. Subsequent analysis and/or sequential manual Edman degradation of the purified phosphopeptides revealed that Thr-65 and Ser-115 were most extensively phophorylated by PKA and Ser-55 by PKC. For the dephosphorylation kinetics, the phosphorylated MBP was treated with calcineurin or PP2C with various time intervals and the reaction was terminated by direct tryptic digest. Both Thr-65 and Ser-115 residues were dephosphorylated more rapidly than any other ones by phosphatases. However it can be differentiated further by first-order kinetics that the PP2B dephosphorylated both Thr-65 and Ser-115 with almost same manner, whereas PP2C dephosphorylated somewhat preferentially the Ser-115.

• BMB Reports
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• v.43 no.4
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• pp.263-267
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• 2010

To characterize the biochemical properties of the PP2A regulatory B subunit, PPP2R5D, we analyzed its phosphorylation sites, stoichiometry and effect on holoenzyme activity. PPP2R5D was phosphorylated on Ser-53, Ser-68, Ser-81, and Ser-566 by protein kinase A, and mutations at all four of these sites abolished any significant phosphorylation in vitro. In HEK293 cells, however, the Ser-566 was the major phosphorylation site after PKA activation by forskolin, with marginal phosphorylation on Ser-81. Inhibitory tyrosine phosphorylation on Tyr-307 of the PP2A catalytic C subunit was decreased after forskolin treatment. Kinetic analysis showed that overall PP2A activity was increased with phosphorylation by PPP2R5D phosphorylation. The apparent Km was reduced from $11.25\;{\mu}M$ to $1.175\;{\mu}M$ with PPP2R5D phosphorylation, resulting in an increase in catalytic activity. These data suggest that PKA-mediated activation of PP2A is enabled by PPP2R5D phosphorylation, which modulates the affinity of the PP2A holoenzyme to its physiological substrates.

• The Korean Journal of Physiology and Pharmacology
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• v.18 no.2
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• pp.155-161
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• 2014

Overexpression of amyloid precursor protein with the Swedish mutation causes abnormal hyperphosphorylation of the microtubule-associated protein tau. Hyperphosphorylated isoforms of tau are major components of neurofibrillary tangles, which are histopathological hallmarks of Alzheimer's disease. Protein phosphatase 2A (PP2A), a major tau protein phosphatase, consists of a structural A subunit, catalytic C subunit, and a variety of regulatory B subunits. The B subunits have been reported to modulate function of the PP2A holoenzyme by regulating substrate binding, enzyme activity, and subcellular localization. In the current study, we characterized regulatory B subunit-specific regulation of tau protein phosphorylation. We showed that the PP2A B subunit PPP2R2A mediated dephosphorylation of tau protein at Ser-199, Ser-202/Thr-205, Thr-231, Ser-262, and Ser-422. Down-regulation of PPP2R5D expression decreased tau phosphorylation at Ser-202/Thr-205, Thr-231, and Ser-422, which indicates activation of the tau kinase glycogen synthase kinase 3 beta ($GSK3{\beta}$) by PP2A with PPP2R5D subunit. The level of activating phosphorylation of the $GSK3{\beta}$ kinase Akt at Thr-308 and Ser-473 were both increased by PPP2R5D knockdown. We also characterized B subunit-specific phosphorylation sites in tau using mass spectrometric analysis. Liquid chromatography-mass spectrometry revealed that the phosphorylation status of the tau protein may be affected by PP2A, depending on the specific B subunits. These studies further our understanding of the function of various B subunits in mediating site-specific regulation of tau protein phosphorylation.

• BMB Reports
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• v.28 no.4
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• pp.331-335
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• 1995

Simian virus 40 (SV40) small-t antigen (small-t) has been known to regulate the activity of a cellular enzyme, protein phosphatase 2A (PP2A), composed of A. B, and C subunits, via binding to the A subunit In the study presented here, the stoichiometry of the binding of small-t to PP2A was determined to be 1: 1. It was also shown that small-t binds to the AC form of PP2A with a higher apparent affinity than it binds to the free A subunit. We also characterized the interaction of PP2A with wild-type and various mutant small-ts. A single-point mutant (Val134Met) and a double-point mutant (Trp147Gly;Leu152 Pro) of small-t exhibited 3-fold and 5-fold lower potencies in inhibiting PP2A activity. respectively. This suggests that the region around amino acids between 134 and 152 of small-t might be important in regulating the enzyme activity of PP2A.

• Toxicological Research
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• v.34 no.4
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• pp.333-341
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• 2018

Ferulate is a phenolic compound abundant in wheat germ and bran and has been investigated for its beneficial activities. The aim of the present study is to evaluate the efficacy of ferulate against the oxidative stress-induced imbalance of protein tyrosine kinases (PTKs), protein tyrosine phosphatases (PTPs), and serine/threonine protein phosphatase 2A (PP2A), in connection with our previous finding that oxidative stress-induced imbalance of PTKs and PTPs is linked with proinflammatory nuclear factor-kappa B $(NF-{\kappa}B)$ activation. To test the effects of ferulate on this process, we utilized two oxidative stress-induced inflammatory models. First, YPEN-1 cells were pretreated with ferulate for 1 hr prior to the administration of 2,2'-Azobis(2-methylpropionamidine) dihydrochloride (AAPH). Second, 20-month-old Sprague-Dawley rats were fed ferulate for 10 days. After ferulate treatment, the activities of PTKs, PTPs, and PP2A were measured because these proteins either directly or indirectly promote $NF-{\kappa}B$ activation. Our results revealed that in YPEN-1 cells, ferulate effectively suppressed AAPH-induced increases in reactive oxygen species (ROS) and $NF-{\kappa}B$ activity, as well as AAPH-induced PTK activation. Furthermore, ferulate also inhibited AAPH-induced PTP and PP2A inactivation. In the aged kidney model, ferulate suppressed aging-induced activation of PTKs and ameliorated aging-induced inactivation of PTPs and PP2A. Thus, herein we demonstrated that ferulate could modulate PTK/PTP balance against oxidative stress-induced inactivation of PTPs and PP2A, which is closely linked with $NF-{\kappa}B$ activation. Based on these results, the ability of ferulate to modulate oxidative stress-related inflammatory processes is established, which suggests that this compound could act as a novel therapeutic agent.

• The Journal of Pediatrics of Korean Medicine
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• v.14 no.2
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• pp.33-46
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• 2000

It has long been known that Jahage(紫河車) extracts of Placenta hominis are effective for immunological and vascular diseases in human body and thus, was used a major constituent of traditional oriental medicines. From full-term human placenta, we have purified a new type anticoagulant protein, PP27, using different chromatographic techniques of a phenyl TSK gel 650M column, DEAE, HA and Mono-Q columns. PP27 showed single band on SDS-PAGE with a molecular mass (Mr) of 27 kDa under denaturing conditions and a calibrated Sepharose 4B column chromatography indicated a molecular mass of 23 kDa, indicating that the value is similar to those of other PP4 enzyme reported to date. Isoelectric point of PP27 was p15.2. The protein was found to inhibit the coagulation time in a concentration-dependent manner. PP27 was acted as a vascular anticoagulant of annexin type, inhibits the blood clotting process by binding of the essential lipids in a reaction which is dependent on $Ca2^+$ ions. In the presence of $Ca2^+$ ions, PP27 combines with platelet membranes neutralizing their procoagulant effect. Coagulation triggered by the addition of thromboplastin/ lipid- mixtures is extinguished by PP27.

• Journal of the Korea Academia-Industrial cooperation Society
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• v.19 no.7
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• pp.95-104
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• 2018

The structures and energies of the anhydrate and hydrate (hydrate rate: h of 1) states of L-alanine (LA) and glycine (G) were calculated by quantum chemical calculations (QCCs) using B3LYP/6-31G(d,p) for four types of conformers (${\beta}$-extended: ${\Phi}/{\Psi}=t-/t+$, $PP_{II}$: g-/t+, $PP_{II}$-like: g-/g+, and ${\alpha}$-helix: g-/g-). In LA and G, which have an imino proton (NH), three conformation types of ${\beta}$-extended, $PP_{II}$-like, and ${\alpha}$-helix were obtained, and water molecules were inserted mainly between the intra-molecular hydrogen bond of $CO{\cdots}HN$ in $PP_{II}$-like and ${\alpha}$-helix, and attached to the CO group in ${\beta}$-extended. In LA and G, $PP_{II}$-like conformers were most stable in the anhydrate and hydrate states, and the result for LA was different from some experimental and theoretical results from other studies reporting that the main stable conformation of alanine oligopeptide was $PP_{II}$. The formation pattern and stability of the conformation of the oligopeptide was strongly dominated by the presence/absence of intra-molecular hydrogen bonding of $CO{\cdots}HN$, or the presence/absence of an $NH_2$ group in the starting amino acid.

• Applied Chemistry for Engineering
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• v.10 no.1
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• pp.46-50
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• 1999

Effects of $(NH_4)_2HPO_4$ and boron compounds ($Na_2B_4O_7:H_3BO_3=5:1\;ratio$) on the flame retarding characteristics of the surface modified wood powder-filled polypropylene composites were studied experimentally. The mechanical properties of m-phenylene dimaleimide(PDMI)-modified polypropylene composite were also compared with those of unmodified one. The flame retardance of $(NH_4)_2HPO_4$-modified wood powder composites was more improved than that of boron compounds-modified one. The impact strength of composites increased and the tensile strength of those decreased progressively with an increase of wood powder loading. The mechanical properties of modified polypropylene composites are more improved than those of unmodified one. The tensile strength also increased marginally with increasing the concentration of flame retardants, yielding a maximum when the concentration of flame retardant is 25.0 wt %. And the tensile strength of the composite was increased up to 16 wt % with increasing concentration of PDMI.