Journal Article

Lipid-targeting pleckstrin homology domain turns its autoinhibitory face toward the TEC kinases

Wales, Thomas E., Amatya, Neha, Yeung, Wayland, Engen, John R., Andreotti, Amy H., Kwon, Annie, Joseph, Raji E., Kannan, Natarajan, Fulton, D. Bruce

2019

Overview

The pleckstrin homology (PH) domain is well known for its phospholipid targeting function. The PH-TEC homology (PHTH) domain within the TEC family of tyrosine kinases is also a crucial component of the autoinhibitory apparatus. The autoinhibitory surface on the PHTH domain has been previously defined, and biochemical investigations have shown that PHTH-mediated inhibition is mutually exclusive with phosphatidylinositol binding. Here we use hydrogen/deuterium exchange mass spectrometry, nuclear magnetic resonance (NMR), and evolutionary sequence comparisons to map where and how the PHTH domain affects the Bruton’s tyrosine kinase (BTK) domain. The data map a PHTH-binding site on the activation loop face of the kinase C lobe, suggesting that the PHTH domain masks the activation loop and the substrate-docking site. Moreover, localized NMR spectral changes are observed for non–surface-exposed residues in the active site and on the distal side of the kinase domain. These data suggest that the association of PHTH induces allosteric conformational shifts in regions of the kinase domain that are critical for catalysis. Through statistical comparisons of diverse tyrosine kinase sequences, we identify residues unique to BTK that coincide with the experimentally determined PHTH-binding surface on the kinase domain. Our data provide a more complete picture of the autoinhibitory conformation adopted by full-length TEC kinases, creating opportunities to target the regulatory domains to control the function of these kinases in a biological setting.

• Publisher: National Academy of Sciences
• Subject: Activation / Agammaglobulinaemia Tyrosine Kinase - chemistry / Agammaglobulinaemia Tyrosine Kinase - genetics / Agammaglobulinaemia Tyrosine Kinase - metabolism / Allosteric properties / Allosteric Regulation / Binding Sites / Biological Sciences / Biophysics and Computational Biology / Bruton's tyrosine kinase / Catalysis / Conformation / Deuterium / Domains / Homology / Humans / Kinases / Lipid Metabolism / Lipids / Lipids - chemistry / Magnetic Resonance Spectroscopy / Masks / Mass spectrometry / Mass spectroscopy / Models, Molecular / NMR / Nuclear magnetic resonance / Phosphatidylinositol / Phospholipids / Pleckstrin / Pleckstrin Homology Domains / Protein Domains / Protein-tyrosine kinase / Protein-Tyrosine Kinases - chemistry / Protein-Tyrosine Kinases - genetics / Protein-Tyrosine Kinases - metabolism / Residues / Substrates / Tyrosine