• The Korean Journal of Physiology and Pharmacology
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• 제2권1호
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• pp.41-48
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• 1998

The present study was undertaken to characterize homocysteic acid (HCA)-and cysteic acid (CA)-mediated formation of inositol phosphates (InsP) in primary culture of rat cerebellar granule cells. HCA and CA stimulated InsP formation in a dose-dependent manner, which was prevented by the N-methyl-D-aspartate (NMDA) receptor antagonist D,L-2-amino-5-phosphopentanoic acid (APV). CA-, but not HCA-, mediated InsP formation was in part prevented by the metabotropic glutamate receptor antagonist ?${\alpha}$-methyl-4-carboxyphenylglycine ($({\pm})$-MCPG). Both HCA- and CA-mediated increases in intracellular calcium concentration were completely blocked by APV, but were not altered by $({\pm})$-MCPG. CA-mediated InsP formation was in part prevented by removal of endogenous glutamate. In contrast, the glutamate transport blocker L-aspartic acid-${\beta}$-hydroxamate synergistically increased CA responses. These data indicate that in cerebellar granule cells HCA mediates InsP formation wholly by activating NMDA receptor. In contrast, CA stimulates InsP formation by activating both NMDA receptor and metabotropic glutamate receptor, and in part by releasing endogenous glutamate into extracellular milieu.

• Molecules and Cells
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• 제40권5호
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• pp.315-321
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• 2017

The inositol polyphosphates are a group of multifunctional signaling metabolites whose synthesis is catalyzed by a family of inositol kinases that are evolutionarily conserved from yeast to humans. Inositol polyphosphate multikinase (IPMK) was first identified as a subunit of the arginine-responsive transcription complex in budding yeast. In addition to its role in the production of inositol tetrakis- and pentakisphosphates ($IP_4$ and $IP_5$), IPMK also exhibits phosphatidylinositol 3-kinase (PI3-kinase) activity. Through its PI3-kinase activity, IPMK activates Akt/PKB and its downstream signaling pathways. IPMK also regulates several protein targets non-catalytically via protein-protein interactions. These non-catalytic targets include cytosolic signaling factors and transcription factors in the nucleus. In this review, we highlight the many known functions of mammalian IPMK in controlling cellular signaling networks and discuss future challenges related to clarifying the unknown roles IPMK plays in physiology and disease.

• 한국미생물·생명공학회지
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• 제25권6호
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• pp.592-597
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• 1997

During the screening of inhibitors against phospholipase C (PLC) and the formation of inositol phosphates (IP$_{t}$) at NIH3T3${\gamma}$1 cells from microbial secondary metabolites, we selected a fungal strain MT60109 which was capable of producing an inhibitor. By the taxonomic studies, this fungus was identified as Pseudallescheria sp. MT60109 and an inhibitor of PLC was purified by BuOH extraction and chromatographic techniques from the culture broth of Pseudallescheria sp. MT60109. The inhibitor was identified as thielavin B by the physico-chemical properties and spectroscopic analysis of UV, FAB-MS, $^{1}$H, $^{13}$C-NMR, $^{1}$H-$^{1}$H COSY and HMBC. Thielavin B showed potent inhibitory activity against PLC purified from bovine brain with an IC$_{50}$ of 20 $\mu$M. And it also inhibited the formation of inositol phosphates in platelet-derived growth factor (PDGF) -stimulated NIH3T3${\gamma}$1 cells with an IC$_{50}$ of 20 $\mu$M.

• 약학회지
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• 제41권4호
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• pp.433-438
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• 1997

MT 51005 as a potent inhibitor of phospholipase C(PLC) was purified from the culture broth of a fungal strain No. 51005 isolated from soil. It was identified as a benzaldehyde d erivative, anguillosporal. by the physico-chemical properties and spectroscopic data. Anguillosporal showed the inhibitory activity against purified PLC with an $IC_{50}\;of\;13{\mu}g/ml$. And it also inhibited the formation of inositol phosphates($IP_t$) in platelet-derived growth factor(PDGF)-stimulated $NIH3T3{\gamma}1$ cells with an $IC_{50}\;of\;0.8{\mu}g/ml$.

• Mycobiology
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• 제28권3호
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• pp.119-122
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• 2000

Phytases (myo-inositol hexakisphosphate phosphohydrolase; EC 3.1.3.8) are enzymes which catalyze the hydrolisys of phytate into myo-inositol and inorganic phosphates. Phytases are found in plants and a variety of microorganisms. Aspergillus species were treated with 254 nm of UV irradiation for the screening of phytase overproducing mutant strains. At 15 minute irradiation, the survivals of population were less than 5%, and UV irradiation time was decided at 20 minute for the isolation of mutant strains. Four UV mutant strains in A. oryzae (YUV-47, -169, -341, -511) and six in A. ficuum (FUV-17, -36, -69, -193, -317, -419) were isolated on PSM media containing ammonium phosphate. The specific enzyme activities of A. ficuum mutants are 110 to 140% higher than that of wild type.

• 약학회지
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• 제44권2호
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• pp.162-168
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• 2000

To investigate the effects of protein kinases on endothelin-1-induced phospholipase C activation and $Ca^{2+}$ mobilization in Rat-2 fibroblast, we measured the formation of inositol phosphates and intracellular $Ca^{2+}$ concentration with [$^3$H]inositol and Fura-2/AM, respectively. Endothelin-1 dose-dependently activated phospholipase C and increased intracellular $Ca^{2+}$ concentration. Protein kinase C activator PMA, significantly inhibited both phospholipase C activity and $Ca^{2+}$ mobilization induced by endothelin-1. Tyrosine kinase inhibitor, genistein, inhibited both. On the other hand, cyclic nucleotide (cAMP and cGMP) did not have any influence on the signaling pathway of phospholipase C-Ca$^{2+}$ mobilization induced by endothelin-1. These results suggest that protein kinase C and tyrosine kinase counteract on the signaling pathway of phospholipase C-Ca$^{2+}$ mobilization induced by endothelin-1 in Rat-2 fibroblast. fibroblast.

• Molecules and Cells
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• 제43권3호
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• pp.222-227
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• 2020

Inositol polyphosphate multikinase (IPMK) is required for the biosynthesis of inositol phosphates (IPs) through the phosphorylation of multiple IP metabolites such as IP3 and IP4. The biological significance of IPMK's catalytic actions to regulate cellular signaling events such as growth and metabolism has been studied extensively. However, pharmacological reagents that inhibit IPMK have not yet been identified. We employed a structure-based virtual screening of publicly available U.S. Food and Drug Administration-approved drugs and chemicals that identified the antidepressant, vilazodone, as an IPMK inhibitor. Docking simulations and pharmacophore analyses showed that vilazodone has a higher affinity for the ATP-binding catalytic region of IPMK than ATP and we validated that vilazodone inhibits IPMK's IP kinase activities in vitro. The incubation of vilazodone with NIH3T3-L1 fibroblasts reduced cellular levels of IP5 and other highly phosphorylated IPs without influencing IP4 levels. We further found decreased Akt phosphorylation in vilazodone-treated HCT116 cancer cells. These data clearly indicate selective cellular actions of vilazodone against IPMK-dependent catalytic steps in IP metabolism and Akt activation. Collectively, our data demonstrate vilazodone as a method to inhibit cellular IPMK, providing a valuable pharmacological agent to study and target the biological and pathological processes governed by IPMK.

• Molecules and Cells
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• 제19권3호
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• pp.375-381
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• 2005

Phospholipase C-${\beta}$ (PLC-${\beta}$) hydrolyses phosphatidylinositol 4,5-bisphosphate and generates inositol 1,4,5-trisphosphate in response to activation of various G protein-coupled receptors (GPCRs). Using glial cells from knock-out mice lacking either PLC-${\beta}1$ [PLC-${\beta}1$ (-/-)] or PLC-${\beta}3$ [PLC-${\beta}3$ (-/-)], we examined which isotype of PLC-${\beta}$ participated in the cellular signaling events triggered by thrombin. Generation of inositol phosphates (IPs) was enhanced by thrombin in PLC-${\beta}1$ (-/-) cells, but was negligible in PLC-${\beta}3$ (-/-) cells. Expression of PLC-${\beta}3$ in PLC-${\beta}3$ (-/-) cells resulted in an increase in pertussis toxin (PTx)-sensitive IPs in response to thrombin as well as to PAR1-specific peptide, while expression of PLC-${\beta}1$ in PLC-${\beta}1$ (-/-) cells did not have any effect on IP generation. The thrombin-induced $[Ca^{2+}]_i$ increase was delayed and attenuated in PLC-${\beta}3$ (-/-) cells, but normal in PLC-${\beta}1$ (-/-) cells. Pertussis toxin evoked a delayed $[Ca^{2+}]_i$ increase in PLC-${\beta}3$ (-/-) cells as well as in PLC-${\beta}1$ (-/-) cells. These results suggest that activation of PLC-${\beta}3$ by pertussis toxin-sensitive G proteins is responsible for the transient $[Ca^{2+}]_i$ increase in response to thrombin, whereas the delayed $[Ca^{2+}]_i$ increase may be due to activation of some other PLC, such as PLC-${\beta}4$, acting via PTx-insensitive G proteins.

• 한국식품위생안전성학회지
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• 제22권1호
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• pp.37-44
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• 2007

피틴산(Phytic acid)dms inositol hexaphosphate로서 식물성 식품 및 씨 중에 1-5%가량 존재하는 자연 항산화 물질로서 소화효소에 의해 쉽게 분해되지 않는다. 한편 인산염은 식육에서 품질개량제로 식육에서의 항균 효과에 대해서는 이미 여러 보고가 있으나 그 결과는 다양하다. 일반적으로 생육에서는 그 효능이 낮은데 이것은 생육에 존재하는 phosphatase의 작용에 의해 인산염들이 가수분해되기 때문이다. 한편 피틴산염은 열에 강하고 쉽게 분해되지 않으므로 생육이나 멸균 처리된 식육에서 공히 인산염들이 가지고 있는 항균효과를 기대할 수 있다. 본 실험에서는 선택배지에서 E. coli O157:H7에 대한 sodium phytate(STP), sodium pyrophosphate(SPP) 및 sodium tripolyphosphate(STPP)의 항균효과를 평가하였고,더불어 생육 및 가열 처리된 식육모델에서 E. coli O157:H7 접종 후에 항균 효과를 비교 평가하였다. 선택된 배지인 tryptic soy broth에서 E. coli O157:H7에 대한 항균 효과는 SPT, SPP 및 STPP의 0.05, 0.1, 0.5%의 세 가지 농도에서 농도에 의존적으로 유의성 있게 세균 증식을 억제시켰으며, 항균효과는 SPP에서 가장 강하게 나타났다. 생육인 닭고기, 돼지고기 및 소고기에 SPT, SPP 및 STPP를 각각 0.05, 0.1, 0.3 그리고 0.5%의 농도별로 첨가했을 때 E. coli O157:H7의 증식이 모두 유의성 있게 감소하였다 (p<0.05). 또한 가열 처리한 식육에서도 이러한 항균효과가 유의적으로 관찰되었으며(p<0.05), SPT는 SPP나 STPP보다 더 강한 항균효과를 보였다. 이러한 항균효과들은 가열처리된 식육보다 생육에서 더욱 강하게 나타났다. SPT, SPP 및 STPP의 첨가는 식육에서 pH를 증가시켰으며, STP의 첨가는 식육에서 soluble orthophosphate 유리에 영향을 주지 않았으나, SPP 및 STPP의 첨가는 식육에서 soluble orthophosphate의 유리를 증가시켰다. 이러한 결과로부터 피틴산염은 이미 알려진 인산염들보다 미약하나마 항균효과가 뛰어나며 더불어 축산식품의 첨가제로서 사용시 식품의 기능성 향상과 더불어 매우 유용하게 사용될 수 있을 것이다.

• 한국식품영양학회지
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• 제6권1호
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• pp.37-46
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• 1993

Brain, heart, liver, lung, kidney and thymus etc. 12 organs were removed and homogenized from Dawley-Sprague rats after suffocation. After fractionation of the tissue cytosols, enzymatic activities of the key enzymes in metabolic inositol phosphates cycle, PLC, IPSK and Ins(1,4,5) P35-phosphatase, were measured respectively. Hybridoma monoclones producing anti-lP3K murine monoclonal antibodies were obtained by the fusion of SP2/Ag 0-14 and spleen cells of mouse immunized with purified 53KDa IPSK, screening and cloning procedures. 18 cloned hybridoma cells were obtained, background due to nonspecific binding was very low with 10 clones. These Abs were purified from ascitic fluids by using affi-gel 15, and determined subtype of Abs. When immunoreactivities for rat tissues IP3K were exercised by adding the mixed Abs of 19Gl and 19G2b, they showed an overall similarity with noncompetitive inhibition. Brain tissue has high sensitivity for anti-lP3K Ab, whereas heart tissue has very low activity. In kinetic parameters Km value was 1.58 mM and Vmx value was 5.41umol/min/ml, respectively Only one form of 40 KDa IPSK was detected in heart tissues, however rat brain contains at least three immunologically distinct IP3K (53, 51 and 40 KDa) in western blot analysis. Of them 53 KDa protein was major enzyme in enzymatic activity. Northern blot analysis with 32P-labeled CDNA probe which encodes 1.8 Kb IPSK gene was performed. These results suggest that IPSK are regulated at transcriptional level during rat tissue development.