| Oxidation of the diphosphoinositol polyphosphate phosphohydrolase-like Nudix hydrolase Aps from Drosophila melanogaster induces thermolability--A possible regulatory switch? | |
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MedLine Citation:
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PMID: 20394834 Owner: NLM Status: MEDLINE |
Abstract/OtherAbstract:
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Unlike mammalian cells, Drosophila melanogaster contains only a single member of the diphosphoinositol polyphosphate phosphohydrolase subfamily of the Nudix hydrolases, suggesting that functional specialisation has not occurred in this organism. In order to evaluate its function, Aps was cloned and characterized. It hydrolyses a range of (di)nucleoside polyphosphates, the most efficient being guanosine 5'-tetraphosphate (K(m)=11 microM, k(cat)=0.79 s(-1)). However, it shows a 5-fold preference for the hydrolysis of diphosphoinositol pentakisphosphate (PP-InsP(5), K(m)=0.07 microM, k(cat)=0.024 s(-1)). Assayed at 26 degrees C, Aps had an alkaline pH optimum and required a divalent ion: Mg(2+) (10-20 mM) or Mn(2+) (1 mM) were preferred for nucleotide hydrolysis and Mg(2+) (0.5-1 mM) or Co(2+) (1-100 microM) for PP-InsP(5) hydrolysis. GFP-fusions showed that Aps was predominantly cytoplasmic, with some nuclear localization. In the absence of dithiothreitol Aps was heat labile, rapidly losing activity even at 36 degrees C, while in the presence of dithiothreitol, Aps was heat stable, surviving for 5 min at 76 degrees C. Heat lability was restored by H(2)O(2) and mass spectrometric analysis suggested that this was due to reversible dimerisation involving two inter-molecular disulphides between Cys23 and Cys25. Aps expression was highest in embryos and declined throughout development. The ratio of PP-InsP(5) to inositol hexakisphosphate also decreased throughout development, with the highest level of PP-InsP(5) found in embryos. These data suggest that the redox state of Aps may play a role in controlling its activity by altering its stability, something that could be important for regulating PP-InsP(5) during development. |
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Authors:
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Lucinda Winward; William G F Whitfield; Alexander G McLennan; Stephen T Safrany |
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Publication Detail:
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Type: Journal Article; Research Support, Non-U.S. Gov't Date: 2010-04-13 |
Journal Detail:
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Title: The international journal of biochemistry & cell biology Volume: 42 ISSN: 1878-5875 ISO Abbreviation: Int. J. Biochem. Cell Biol. Publication Date: 2010 Jul |
Date Detail:
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Created Date: 2010-06-02 Completed Date: 2010-09-27 Revised Date: - |
Medline Journal Info:
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Nlm Unique ID: 9508482 Medline TA: Int J Biochem Cell Biol Country: Netherlands |
Other Details:
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Languages: eng Pagination: 1174-81 Citation Subset: IM |
Copyright Information:
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Copyright 2010 Elsevier Ltd. All rights reserved. |
Affiliation:
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Division of Cell Signalling, University of Dundee, Dow Street, Dundee DD1 4HN, United Kingdom. lusi83@hotmail.com |
Export Citation:
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APA/MLA Format Download EndNote Download BibTex |
| MeSH Terms | |
Descriptor/Qualifier:
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Acid Anhydride Hydrolases
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chemistry,
genetics,
metabolism* Adenine Nucleotides / metabolism Amino Acid Sequence Animals Cations, Divalent / pharmacology Chromatography, High Pressure Liquid Dithiothreitol / pharmacology Drosophila Proteins / chemistry, genetics, metabolism* Drosophila melanogaster / enzymology*, genetics Enzyme Stability / drug effects Gene Expression Regulation, Developmental / drug effects Hydrogen-Ion Concentration / drug effects Hydrolysis / drug effects Inositol Phosphates / metabolism Kinetics Molecular Sequence Data Oxidation-Reduction / drug effects Protein Transport / drug effects Pyrophosphatases / chemistry, genetics, metabolism* Sequence Alignment Subcellular Fractions / drug effects, enzymology Substrate Specificity / drug effects Temperature* |
| Grant Support | |
ID/Acronym/Agency:
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//Biotechnology and Biological Sciences Research Council |
| Chemical | |
Reg. No./Substance:
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0/Adenine Nucleotides; 0/Cations, Divalent; 0/Drosophila Proteins; 0/Inositol Phosphates; 148077-18-3/1-diphosphoinositol pentakisphosphate; 3483-12-3/Dithiothreitol; EC 3.6.-/Acid Anhydride Hydrolases; EC 3.6.1.-/Aps protein, Drosophila; EC 3.6.1.-/Pyrophosphatases; EC 3.6.1.-/diphosphoinositol polyphosphate phosphohydrolase; EC 3.6.1.-/nudix hydrolases |
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine
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