5c46
Crystal structure of an engineered construct of phosphatidylinositol 4 kinase III beta in complex with GTP gamma S loaded Rab11
Structural highlights
FunctionPI4KB_HUMAN Phosphorylates phosphatidylinositol (PI) in the first committed step in the production of the second messenger inositol-1,4,5,-trisphosphate (PIP). May regulate Golgi disintegration/reorganization during mitosis, possibly via its phosphorylation. Involved in Golgi-to-plasma membrane trafficking (By similarity).[1] [2] [3] Publication Abstract from PubMedThe ability of proteins to bind and interact with protein partners plays fundamental roles in many cellular contexts. X-ray crystallography has been a powerful approach to understand protein-protein interactions, however, a challenge in the crystallization of proteins and their complexes is the presence of intrinsically disordered regions. In this article we describe an application of hydrogen deuterium exchange mass spectrometry (HDX-MS) to identify dynamic regions within type III phosphatidylinositol 4 kinase beta (PI4KIIIbeta) in complex with the GTPase Rab11. This information was then used to design deletions that allowed for the production of diffraction quality crystals. Importantly we also used HDX-MS to verify that the new construct was properly folded, consistent with it being catalytically and functionally active. Structures of PI4KIIIbeta in an Apo state and bound to the potent inhibitor BQR695 in complex with both GTPgammaS and GDP loaded Rab11 were determined. This hybrid HDX-MS/crystallographic strategy revealed novel aspects of the PI4KIIIbeta-Rab11 complex, as well as the molecular mechanism of potency of a PI4K specific inhibitor (BQR695). This approach is widely applicable to protein-protein complexes, and is an excellent strategy to optimize constructs for high-resolution structural approaches. This article is protected by copyright. All rights reserved. Using hydrogen deuterium exchange mass spectrometry to engineer optimized constructs for crystallization of protein complexes: Case study of PI4KIIIbeta with Rab11.,Fowler ML, McPhail JA, Jenkins ML, Masson GR, Rutaganira FU, Shokat KM, Williams RL, Burke JE Protein Sci. 2016 Jan 12. doi: 10.1002/pro.2879. PMID:26756197[4] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. See AlsoReferences
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