Crystallographic and kinetic analyses of human IPMK reveal disordered domains modulate ATP binding and kinase activity

Seacrist, Corey D.; Blind, Raymond D.

doi:10.1038/s41598-018-34941-3

Title: Crystallographic and kinetic analyses of human IPMK reveal disordered domains modulate ATP binding and kinase activity

Journal Article · 11 November 2018 · Scientific Reports

DOI: https://doi.org/10.1038/s41598-018-34941-3 · OSTI ID:1544877

Seacrist, Corey D. ^[1];

^[1]

Vanderbilt Univ. School of Medicine, Nashville, TN (United States)

Inositol polyphosphate multikinase (IPMK) is a member of the IPK-superfamily of kinases, catalyzing phosphorylation of several soluble inositols and the signaling phospholipid PI(4,5)P₂ (PIP₂). IPMK also has critical non-catalytic roles in p53, mTOR/Raptor, TRAF6 and AMPK signaling mediated partly by two disordered domains. Although IPMK non-catalytic functions are well established, it is less clear if the disordered domains are important for IPMK kinase activity or ATP binding. Here, kinetic and structural analyses of an engineered human IPMK lacking all disordered domains (ΔIPMK) are presented. Although the K_M for PIP₂ is identical between ΔIPMK and wild type, ΔIPMK has a 1.8-fold increase in k_cat for PIP₂, indicating the native IPMK disordered domains decrease IPMK activity in vitro. The 2.5 Å crystal structure of ΔIPMK is reported, confirming the conserved ATP-grasp fold. A comparison with other IPK-superfamily structures revealed a putative “ATP-clamp” in the disordered N-terminus, we predicted would stabilize ATP binding. Consistent with this observation, removal of the ATP clamp sequence increases the K_M for ATP 4.9-fold, indicating the N-terminus enhances ATP binding to IPMK. Together, these structural and kinetic studies suggest in addition to mediating protein-protein interactions, the disordered domains of IPMK impart modulatory capacity to IPMK kinase activity through multiple kinetic mechanisms.

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Research Organization:: Argonne National Laboratory (ANL), Argonne, IL (United States). Advanced Photon Source (APS)

Sponsoring Organization:: USDOE Office of Science (SC)

OSTI ID:: 1544877

Journal Information:: Scientific Reports, Journal Name: Scientific Reports Journal Issue: 1 Vol. 8; ISSN 2045-2322

Publisher:: Nature Publishing GroupCopyright Statement

Country of Publication:: United States

Language:: ENGLISH

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Synthesis of diphosphoinositol pentakisphosphate by a newly identified family of higher inositol polyphosphate kinases Saiardi, Adolfo; Erdjument-Bromage, Hediye; Snowman, Adele M. Current Biology, Vol. 9, Issue 22 https://doi.org/10.1016/S0960-9822(00)80055-X	journal	November 1999
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The Lipid Kinase PI5P4Kβ Is an Intracellular GTP Sensor for Metabolism and Tumorigenesis Sumita, Kazutaka; Lo, Yu-Hua; Takeuchi, Koh Molecular Cell, Vol. 61, Issue 2 https://doi.org/10.1016/j.molcel.2015.12.011	journal	January 2016
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Structural features of human inositol phosphate multikinase rationalize its inositol phosphate kinase and phosphoinositide 3-kinase activities Wang, Huanchen; Shears, Stephen B. Journal of Biological Chemistry, Vol. 292, Issue 44 https://doi.org/10.1074/jbc.M117.801845	journal	September 2017
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Visualization and Measurement of ATP Levels in Living Cells Replicating Hepatitis C Virus Genome RNA Ando, Tomomi; Imamura, Hiromi; Suzuki, Ryosuke PLoS Pathogens, Vol. 8, Issue 3 https://doi.org/10.1371/journal.ppat.1002561	journal	March 2012
Nucleocytoplasmic shuttling of human inositol phosphate multikinase is influenced by CK2 phosphorylation Meyer, Rüdiger; Nalaskowski, Marcus M.; Ehm, Patrick Biological Chemistry, Vol. 393, Issue 3 https://doi.org/10.1515/hsz-2011-0209	journal	March 2012
Inositol Diphosphate Signaling Regulates Telomere Length D., York, John; N., Armbruster, Blaine; Patricia, Greenwell, The University of North Carolina at Chapel Hill University Libraries https://doi.org/10.17615/pgat-ss03	text	January 2005
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