The kinetic properties of a human PPIP5K reveal that its kinase activities are protected against the consequences of a deteriorating cellular bioenergetic environment
| Autor: | Huanchen Wang, Stephen B. Shears, Jeremy D. Weaver |
|---|---|
| Jazyk: | angličtina |
| Rok vydání: | 2013 |
| Předmět: |
InsP8
1 5-bis-diphosphoinositol 2 3 4 6-tetrakisphosphate InsS6 inositol hexasulphate Bioenergetics NCBI National Center for Biotechnology Information InsP5 inositol 1 3 4 5 6-pentakisphosphate Mutant lcsh:Life lcsh:QR1-502 bis-diphosphoinositol tetrakisphosphate 1-InsP7 1-diphosphoinositol 2 3 4 5 6-pentakisphosphate Biochemistry PPIP5K2KD human diphosphoinositol pentakisphosphate kinase 2 kinase domain lcsh:Microbiology Substrate Specificity chemistry.chemical_compound Adenine nucleotide GST glutathione transferase Inositol chemistry.chemical_classification Kinase InsP6 inositol hexakisphosphate cellular energy homoeostasis InsP4 inositol 1 3 4 5-tetrakisphosphate diphosphoinositol polyphosphate Metabolic Networks and Pathways Biotechnology Reaction mechanism Inositol Phosphates Kinetics Biophysics Biology S2 Catalysis Phosphates Genetics Humans Molecular Biology IP6K inositol hexakisphosphate kinase Original Paper PPIP5K diphosphoinositol pentakisphosphate kinase Phosphotransferases (Phosphate Group Acceptor) InsP3 inositol 1 3 4trisphosphate inositol 1 3 4-trisphosphate 5/6-kinase (ITPK1) DIPP diphosphoinositol-polyphosphate phosphohydrolase 5-InsP7 1-diphosphoinositol 2 3 4 5 6-pentakisphosphate Substrate (chemistry) Cell Biology InsP inositol phosphate diphosphoinositol pentakisphosphate PP-InsPs diphosphoinositol polyphosphates lcsh:QH501-531 Enzyme chemistry DTT dithiothreitol inositol pyrophosphate ITPK1 inositol 1 3 4-trisphosphate 5/6-kinase Energy Metabolism |
| Zdroj: | Bioscience Reports, Vol 33, Iss 2, p e00022 (2013) ResearcherID BASE-Bielefeld Academic Search Engine Bioscience Reports |
| ISSN: | 1573-4935 0144-8463 |
| Popis: | We obtained detailed kinetic characteristics–stoichiometry, reaction rates, substrate affinities and equilibrium conditions–of human PPIP5K2 (diphosphoinositol pentakisphosphate kinase 2). This enzyme synthesizes ‘high-energy’ PP-InsPs (diphosphoinositol polyphosphates) by metabolizing InsP6 (inositol hexakisphosphate) and 5-InsP7 (5-diphosphoinositol 1,2,3,4,6-pentakisphosphate) to 1-InsP7 (1-diphosphoinositol 2,3,4,5,6-pentakisphosphate) and InsP8 (1,5-bis-diphosphoinositol 2,3,4,6-tetrakisphosphate), respectively. These data increase our insight into the PPIP5K2 reaction mechanism and clarify the interface between PPIP5K catalytic activities and cellular bioenergetic status. For example, stochiometric analysis uncovered non-productive, substrate-stimulated ATPase activity (thus, approximately 2 and 1.2 ATP molecules are utilized to synthesize each molecule of 1-InsP7 and InsP8, respectively). Impaired ATPase activity of a PPIP5K2-K248A mutant increased atomic-level insight into the enzyme's reaction mechanism. We found PPIP5K2 to be fully reversible as an ATP-synthase in vitro, but our new data contradict previous perceptions that significant ‘reversibility’ occurs in vivo. PPIP5K2 was insensitive to physiological changes in either [AMP] or [ATP]/[ADP] ratios. Those data, together with adenine nucleotide kinetics (ATP Km=20–40 μM), reveal how insulated PPIP5K2 is from cellular bioenergetic challenges. Finally, the specificity constants for PPIP5K2 revise upwards by one-to-two orders of magnitude the inherent catalytic activities of this enzyme, and we show its equilibrium point favours 80–90% depletion of InsP6/5-InsP7. |
| Databáze: | OpenAIRE |
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