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1.
Ohnishi T Ohba H Seo KC Im J Sato Y Iwayama Y Furuichi T Chung SK Yoshikawa T 《The Journal of biological chemistry》2007,282(1):637-646
Lithium is used in the clinical treatment of bipolar disorder, a disease where patients suffer mood swings between mania and depression. Although the mode of action of lithium remains elusive, a putative primary target is thought to be inositol monophosphatase (IMPase) activity. Two IMPase genes have been identified in mammals, the well characterized myo-inositol monophosphatase 1 (IMPA1) and myo-inositol monophosphatase 2 (IMPA2). Several lines of genetic evidence have implicated IMPA2 in the pathogenesis of not only bipolar disorder but also schizophrenia and febrile seizures. However, little is known about the protein, although it is predicted to have lithium-inhibitable IMPase activity based on its homology to IMPA1. Here we present the first biochemical study comparing the enzyme activity of IMPA2 to that of IMPA1. We demonstrate that in vivo, IMPA2 forms homodimers but no heterodimers with IMPA1. Recombinant IMPA2 exhibits IMPase activity, although maximal activity requires higher concentrations of magnesium and a higher pH. IMPA2 shows significantly lower activity toward myo-inositol monophosphate than IMPA1. We therefore screened for additional substrates that could be more efficiently dephosphorylated by IMPA2, but failed to find any. Importantly, when using myo-inositol monophosphate as a substrate, the IMPase activity of IMPA2 was inhibited at high lithium and restricted magnesium concentrations. This kinetics distinguishes it from IMPA1. We also observed a characteristic pattern of differential expression between IMPA1 and IMPA2 in a selection of tissues including the brain, small intestine, and kidney. These data suggest that IMPA2 has a separate function in vivo from that of IMPA1. 相似文献
2.
The structure of the first tetrameric inositol monophosphatase (IMPase) has been solved. This enzyme, from the eubacterium Thermotoga maritima, similarly to its archaeal homologs exhibits dual specificity with both IMPase and fructose-1,6-bisphosphatase activities. The tetrameric structure of this unregulated enzyme is similar, in its quaternary assembly, to the allosterically regulated tetramer of fructose-1,6-bisphosphatase. The individual dimers are similar to the human IMPase. Additionally, the structures of two crystal forms of IMPase show significant differences. In the first crystal form, the tetrameric structure is symmetrical, with the active site loop in each subunit folded into a beta-hairpin conformation. The second form is asymmetrical and shows an unusual structural change. Two of the subunits have the active site loop folded into a beta-hairpin structure, whereas in the remaining two subunits the same loop adopts an alpha-helical conformation. 相似文献
3.
Although neurons are highly polarized, how neuronal polarity is generated remains poorly understood. An evolutionarily conserved inositol-producing enzyme myo-inositol monophosphatase (IMPase) is essential for polarized localization of synaptic molecules in Caenorhabditis elegans and can be inhibited by lithium, a drug for bipolar disorder. The synaptic defect of IMPase mutants causes defects in sensory behaviors including thermotaxis. Here we show that the abnormalities of IMPase mutants can be suppressed by mutations in two enzymes, phospholipase Cβ or synaptojanin, which presumably reduce the level of membrane phosphatidylinositol 4,5-bisphosphate (PIP(2)). We also found that mutations in phospholipase Cβ conferred resistance to lithium treatment. Our results suggest that reduction of PIP(2) on plasma membrane is a major cause of abnormal synaptic polarity in IMPase mutants and provide the first in vivo evidence that lithium impairs neuronal PIP(2) synthesis through inhibition of IMPase. We propose that the PIP(2) signaling regulated by IMPase plays a novel and fundamental role in the synaptic polarity. 相似文献
4.
Myo-inositol (Ins) is a major compatible osmolyte in many cells, including those of Mozambique tilapia (Oreochromis mossambicus). Ins biosynthesis is highly up-regulated in tilapia and other euryhaline fish exposed to hyperosmotic stress. In this study, enzymatic regulation of two enzymes of Ins biosynthesis, Ins phosphate synthase (MIPS) and inositol monophosphatase (IMPase), by direct ionic effects is analyzed. Specific MIPS and IMPase isoforms from Mozambique tilapia (MIPS-160 and IMPase 1) were selected based on experimental, phylogenetic, and structural evidence supporting their role for Ins biosynthesis during hyperosmotic stress. Recombinant tilapia IMPase 1 and MIPS-160 activity was assayed in vitro at ionic conditions that mimic changes in the intracellular milieu during hyperosmotic stress. The in vitro activities of MIPS-160 and IMPase 1 are highest at alkaline pH of 8.8. IMPase 1 catalytic efficiency is strongly increased during hyperosmolality (particularly for the substrate D-Ins-3-phosphate, Ins-3P), mainly as a result of [Na+] elevation. Furthermore, the substrate-specificity of IMPase 1 towards D-Ins-1-phosphate (Ins-1P) is lower than towards Ins-3P. Because MIPS catalysis results in Ins-3P this results represents additional evidence for IMPase 1 being the isoform that mediates Ins biosynthesis in tilapia. Our data collectively demonstrate that the Ins biosynthesis enzymes are activated under ionic conditions that cells are exposed to during hypertonicity, resulting in Ins accumulation, which, in turn, results in restoration of intracellular ion homeostasis. We propose that the unique and direct ionic regulation of the activities of Ins biosynthesis enzymes represents an efficient biochemical feedback loop for regulation of intracellular physiological ion homeostasis during hyperosmotic stress. 相似文献
5.
Navarro-Aviñó JP Bellés JM Serrano R 《Biochemical and biophysical research communications》2003,302(1):41-45
Yeast lithium-sensitive inositol monophosphatase (IMPase) is encoded by a non-essential gene pair (IMP1 and IMP2). Inhibition of IMPase with either Li(+) or Na(+) or a double null mutation imp1 imp2 causes increased levels of inositol monophosphates and reduced level of inositol 1,4,5-trisphosphate. Overexpression of the IMP2 gene has the opposite effects and these results suggest that IMPase activity is limiting for the inositol cycle. Addition of ammonium to cells starved for this nutrient results in a decrease of inositol monophosphates and an increase of inositol 1,4,5-triphosphate, pointing to simultaneous regulation of both inositol 1,4,5-triphosphate production and IMPase activity. 相似文献
6.
Bahn JH Kim AY Jang SH Lee BR Ahn JY Joo HM Kan TC Won MH Kwon HY Kang JH Kwon OS Kim HB Cho SW Lee KS Park J Choi SY 《Molecules and cells》2002,13(1):21-27
Five monoclonal antibodies that recognize porcine brain myo-inositol monophosphate phosphatase (IMPase) have been selected and designated as mAb IMPP 9, IMPP 10, IMPP 11, IMPP 15, and IMPP 17. These antibodies recognize different epitopes of the enzyme and one of these inhibited the enzyme activity. When the total proteins of the porcine brain homogenate separated by SDS-PAGE were probed with monoclonal antibodies, a single reactive protein band of 29 kDa, co-migrating with the purified porcine brain IMPase, was detected. Using the anti-IMPase antibodies as probes, the cross reactivities of the brain IMPase from human and other mammalian tissues, as well as from avian sources, were investigated. Among the human and animal tissues tested, the immunoreactive bands on Western blots appeared to have the same molecular mass of 29 kDa. In addition, there was IMPase immunoreactivity in the various neuronal populations in the rat brain. These results indicate that mammalian brains contain only one major type of immunologically similar IMPase, although some properties of the enzymes that were previously reported differ from each another. The first demonstration of the IMPase localization in the brain may also provide useful data for future investigations on the function of this enzyme in relation to various neurological diseases. 相似文献
7.
Inositol monophosphatase (IMPase) family of proteins are Mg(2+) activated Li(+) inhibited class of ubiquitous enzymes with promiscuous substrate specificity. Herein, the molecular basis of IMPase substrate specificity is delineated by comparative crystal structural analysis of a Staphylococcal dual specific IMPase/NADP(H) phosphatase (SaIMPase - I) with other IMPases of different substrate compatibility, empowered by in silico docking and Escherichia coli SuhB mutagenesis analysis. Unlike its eubacterial and eukaryotic NADP(H) non-hydrolyzing counterparts, the composite structure of SaIMPase - I active site pocket exhibits high structural resemblance with archaeal NADP(H) hydrolyzing dual specific IMPase/FBPase. The large and shallow SaIMPase - I active site cleft efficiently accommodate large incoming substrates like NADP(H), and therefore, justifies the eminent NADP(H) phosphatase activity of SaIMPase - I. Compared to other NADP(H) non-hydrolyzing IMPases, the profound difference in active site topology as well as the unique NADP(H) recognition capability of SaIMPase - I stems from the differential length and orientation of a distant helix α4 (in human and bovine α5) and its preceding loop. We identified the length of α4 and its preceding loop as the most crucial factor that regulates IMPase substrate specificity by employing a size exclusion mechanism. Hence, in SaIMPase - I, the substrate promiscuity is a gain of function by trimming the length of α4 and its preceding loop, compared to other NADP(H) non-hydrolyzing IMPases. This study thus provides a biochemical - structural framework revealing the length and orientation of α4 and its preceding loop as the predisposing factor for the determination of IMPase substrate specificity. 相似文献
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10.
Berggard T Szczepankiewicz O Thulin E Linse S 《The Journal of biological chemistry》2002,277(44):41954-41959
Calbindin D(28k) (calbindin) is a member of the calmodulin superfamily of Ca(2+)-binding proteins. An intracellular target of calbindin was discovered using bacteriophage display. Human recombinant calbindin was immobilized on magnetic beads and used in affinity purification of phage-displayed peptides from a random 12-mer peptide library. One sequence, SYSSIAKYPSHS, was strongly selected both in the presence of Mg(2+) and in the presence of Ca(2+). Homology search against the protein sequence data base identified a closely similar sequence, ISSIKEKYPSHS, at residues 55-66 in myo-inositol-1(or 4)-monophosphatase (IMPase, EC ), which constitute a strongly conserved and exposed region in the three-dimensional structure. IMPase is a key enzyme in the regulation of the activity of the phosphatidylinositol-signaling pathway. It catalyzes the hydrolysis of myo-inositol-1(or 4)-monophosphate to form free myo-inositol, maintaining a supply that represents the precursor for inositol phospholipid second messenger signaling systems. Fluorescence spectroscopy showed that isolated calbindin and IMPase interact with an apparent equilibrium dissociation constant, K(D), of 0.9 microm. Both apo and Ca(2+)-bound calbindin was found to activate IMPase up to 250-fold, depending on the pH and substrate concentration. The activation is most pronounced at conditions that otherwise lead to a very low activity of IMPase, i.e. at reduced pH and at low substrate concentration. 相似文献
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Calbindin-D28k is a calcium binding protein that is highly expressed in the mammalian central nervous system. It has been reported that calbindin-D28k binds to and increases the activity of inositol Monophosphatase (IMPase). This is an enzyme that is involved in the homeostasis of the Inositol trisphosphate signalling cascade by catalysing the final dephosphorylation of inositol and has been implicated in the therapeutic mechanism of lithium treatment of bipolar disorder. Previously studies have shown that calbindin-D28k can increase IMPase activity by up to 250 hundred-fold. A preliminary in silico model was proposed for the interaction.Here, we aimed at exploring the shape and properties of the calbindin-IMPase complex to gain new insights on this biologically important interaction. We created several fusion constructs of calbindin-D28k and IMPase, connected by flexible amino acid linkers of different lengths and orientations to fuse the termini of the two proteins together. The resulting fusion proteins have activities 200%–400% higher the isolated wild-type IMPase. The constructs were characterized by small angle X-ray scattering to gain information on the overall shape of the complexes and validate the previous model. The fusion proteins form a V-shaped, elongated and less compact complex as compared to the model. Our results shed new light into this protein-protein interaction. 相似文献
13.
In sequenced genomes, protein coding regions with unassigned function constitute between 10 and 50% of all open reading frames. Often key enzymes cannot be identified using sequence homology searches. For example, despite the fact that methanogens have an apparently functional gluconeogenesis pathway, standard tools have been unable to identify a fructose-1,6-bisphosphatase (FBPase) gene in the sequenced Methanoccocus jannaschii genome. Using a combination of functional and structural tools, we have shown that the protein product of the M. jannaschii gene MJ0109, which had been tentatively annotated as an inositol monophosphatase (IMPase), has both IMPase and FBPase activities. Moreover, several gene products annotated as IMPases from different thermophilic organisms also possess FBPase activity. Thus, we have found the FBPase that was 'missing' in thermophiles and shown that it also functions as an IMPase. 相似文献
14.
The genome sequence of the cyanobacterium Synechocystis sp. PCC6803 revealed four Open reading frame (ORF) encoding putative inositol monophosphatase or inositol monophosphatase-like
proteins. One of the ORFs, sll1383, is ∼870 base pair long and has been assigned as a probable myo-inositol 1 (or 4) monophosphatase (IMPase; EC 3.1.3.25). IMPase is the second enzyme in the inositol biosynthesis pathway
and catalyses the conversion of L-myo-inositol 1-phosphate to free myo-inositol. The present work describes the functional assignment of ORF sll1383 as myo-inositol 1-phosphate phosphatase (IMPase) through molecular cloning, bacterial overexpression, purification and biochemical
characterization of the gene product. Affinity (K
m) of the recombinant protein for the substrate DL-myo-inositol 1-phosphate was found to be much higher (0.0034 ± 0.0003 mM) compared to IMPase(s) from other sources but in comparison
V
max (∼0.033 μmol Pi/min/mg protein) was low. Li+ was found to be an inhibitor (IC50 6.0 mM) of this enzyme, other monovalent metal ions (e.g. Na+, K+ NH4+) having no significant effect on the enzyme activity. Like other IMPase(s), the activity of this enzyme was found to be totally
Mg2+ dependent, which can be substituted partially by Mn2+. However, unlike other IMPase(s), the enzyme is optimally active at ∼42°C. To the best of our knowledge, sll1383 encoded
IMPase has the highest substrate affinity and specificity amongst the known examples from other prokaryotic sources. A possible
application of this recombinant protein in the enzymatic coupled assay of L-myo-inositol 1-phosphate synthase (MIPS) is discussed. 相似文献
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T Iizasa F Takeuchi Z Honda Y Nishida N Kamatani T Miyamoto 《Biochimica et biophysica acta》1986,882(2):228-233
WI-L2 cells (a B-lymphoblastoid cell line) were more resistant than CEM cells (a T-lymphoblastoid cell line) to deoxyadenosine, ara-A (9-beta-D-arabinofuranosyladenine), or ara-C (1-beta-D-arabinofuranosylcytosine) inhibition. This was caused by a difference in the composition of cytosol 5'-nucleotidases between WI-L2 and CEM cells. In intact cells, the endogenous production of deoxyadenosine from WI-L2 cells deficient in adenosine kinase (EC 2.7.1.20) and deoxycytidine kinase (EC 2.7.1.74) was consistently high, despite changes in endogenous adenosine production. Endogenous production of deoxyadenosine from CEM cells deficient in adenosine kinase and deoxycytidine kinase was, however, coordinated with endogenous adenosine production. In broken cells, cytosol dAMPase (2'-deoxyadenosine 5'-monophosphate 5'-nucleotidase) activity of WI-L2 cells was 3-5-fold higher than that of CEM cells. dAMPase activity could be separated from ATP-activated IMPase (inosine 5'-monophosphate 5'-nucleotidase) by gel filtration (molecular weight: dAMPase; 39,000-46,000; ATP-activated IMPase, greater than 150,000). Cytosol ATP-activated IMPase and dAMPase were isolated by phosphocellulose or DEAE-Bio-Gel A chromatography from non-specific phosphatases. The ATP-activated IMPase showed only marginal activity towards dAMP (2'-deoxyadenosine 5'-monophosphate), ara-AMP (9-beta-D-arabinofuranosyladenine 5'-monophosphate), or ara-CMP (cytosine-beta-D-arabinofuranoside 5'-monophosphate), even in the presence of ATP. The activity of ATP-activated IMPase was similar in WI-L2 and CEM cells. dAMPase was separated into two peaks by DEAE-Bio-Gel A chromatography; one of these peaks degraded ara-AMP and ara-CMP. The activities of both peaks from WI-L2 cells were higher than those from CEM cells. These results show that the degradation of dAMP, ara-AMP or ara-CMP was more specific and rapid in WI-L2 than in CEM cells. 相似文献
17.
Juvenile neuronal ceroid lipofuscinosis (Batten disease) is a neurodegenerative disorder caused by mutation in CLN3. Defective autophagy and concomitant accumulation of autofluorescence enriched with mitochondrial ATP synthase subunit c were previously discovered in Cln3 mutant knock-in mice. In this study, we show that treatment with lithium reduces numbers of LC3-positive autophagosomes and accumulation of LC3-II in Cln3 mutant knock-in cerebellar cells (CbCln3(Δex7/8/Δex7/8) ). Lithium, an inhibitor of GSK3 and IMPase, reduces the accumulation of mitochondrial ATP synthase subunit c and autofluorescence in CbCln3(Δex7/8/Δex7/8) cells, and mitigates the abnormal subcellular distribution of acidic vesicles in the cells. L690,330, an IMPase inhibitor, is as effective as lithium in restoring autophagy in CbCln3(Δex7/8/Δex7/8) cells. Moreover, lithium or down-regulation of IMPase expression protects CbCln3(Δex7/8/Δex7/8) cells from cell death induced by amino acid deprivation. These results suggest that lithium overcomes the autophagic defect in CbCln3(Δex7/8/Δex7/8) cerebellar cells probably through IMPase, thereby reducing their vulnerability to cell death. 相似文献
18.
Alistair K Brown Guoyu Meng Hemza Ghadbane David J Scott Lynn G Dover Jérôme Nigou Gurdyal S Besra Klaus Fütterer 《BMC structural biology》2007,7(1):55
Background
The cell wall of Mycobacterium tuberculosis contains a wide range of phosphatidyl inositol-based glycolipids that play critical structural roles and, in part, govern pathogen-host interactions. Synthesis of phosphatidyl inositol is dependent on free myo-inositol, generated through dephosphorylation of myo-inositol-1-phosphate by inositol monophosphatase (IMPase). Human IMPase, the putative target of lithium therapy, has been studied extensively, but the function of four IMPase-like genes in M. tuberculosis is unclear. 相似文献19.
We recently showed that lithium induces autophagy via inositol monophosphatase (IMPase) inhibition, leading to free inositol depletion and reduced myo-inositol-1,4, 5-triphosphate (IP3) levels. This represents a novel way of regulating mammalian autophagy, independent of the mammalian target of rapamycin (mTOR). Induction of autophagy by lithium led to enhanced clearance of autophagy substrates, like mutant huntingtin fragments and mutant alpha-synucleins, associated with Huntington's disease (HD) and some autosomal dominant forms of Parkinson's disease (PD), respectively. Similar effects were observed with a specific IMPase inhibitor and mood-stabilizing drugs that decrease inositol levels. This may represent a new therapeutic strategy for upregulating autophagy in the treatment of neurodegenerative disorders, where the mutant protein is an autophagy substrate. In this Addendum, we review these findings, and some of the speculative possibilities they raise. 相似文献
20.
Zheng Li Kimberly A. Stieglitz Anthony L. Shrout Yang Wei Robert M. Weis Boguslaw Stec Mary F. Roberts 《Protein science : a publication of the Protein Society》2010,19(2):309-318
The inositol monophosphatase (IMPase) enzyme from the hyperthermophilic archaeon Methanocaldococcus jannaschii requires Mg2+ for activity and binds three to four ions tightly in the absence of ligands: KD = 0.8 μM for one ion with a KD of 38 μM for the other Mg2+ ions. However, the enzyme requires 5–10 mM Mg2+ for optimum catalysis, suggesting substrate alters the metal ion affinity. In crystal structures of this archaeal IMPase with products, one of the three metal ions is coordinated by only one protein contact, Asp38. The importance of this and three other acidic residues in a mobile loop that approaches the active site was probed with mutational studies. Only D38A exhibited an increased kinetic KD for Mg2+; D26A, E39A, and E41A showed no significant change in the Mg2+ requirement for optimal activity. D38A also showed an increased Km, but little effect on kcat. This behavior is consistent with this side chain coordinating the third metal ion in the substrate complex, but with sufficient flexibility in the loop such that other acidic residues could position the Mg2+ in the active site in the absence of Asp38. While lithium ion inhibition of the archaeal IMPase is very poor (IC50~250 mM), the D38A enzyme has a dramatically enhanced sensitivity to Li+ with an IC50 of 12 mM. These results constitute additional evidence for three metal ion assisted catalysis with substrate and product binding reducing affinity of the third necessary metal ion. They also suggest a specific mode of action for lithium inhibition in the IMPase superfamily. 相似文献