title: Carboxyl group footprinting mass spectrometry and molecular dynamics identify key interactions in the HER2-HER3 receptor tyrosine kinase interface.
abstract: The HER2 receptor tyrosine kinase is a driver oncogene in many human cancers, including breast and gastric cancer. Under physiologic levels of expression, HER2 heterodimerizes with other members of the EGF receptor/HER/ErbB family, and the HER2-HER3 dimer forms one of the most potent oncogenic receptor pairs. Previous structural biology studies have individually crystallized the kinase domains of HER2 and HER3, but the HER2-HER3 kinase domain heterodimer structure has yet to be solved. Using a reconstituted membrane system to form HER2-HER3 kinase domain heterodimers and carboxyl group footprinting mass spectrometry, we observed that HER2 and HER3 kinase domains preferentially form asymmetric heterodimers with HER3 and HER2 monomers occupying the donor and acceptor kinase positions, respectively. Conformational changes in the HER2 activation loop, as measured by changes in carboxyl group labeling, required both dimerization and nucleotide binding but did not require activation loop phosphorylation at Tyr-877. Molecular dynamics simulations on HER2-HER3 kinase dimers identify specific inter- and intramolecular interactions and were in good agreement with MS measurements. Specifically, several intermolecular ionic interactions between HER2 Lys-716-HER3 Glu-909, HER2 Glu-717-HER3 Lys-907, and HER2 Asp-871-HER3 Arg-948 were identified by molecular dynamics. We also evaluated the effect of the cancer-associated mutations HER2 D769H/D769Y, HER3 E909G, and HER3 R948K (also numbered HER3 E928G and R967K) on kinase activity in the context of this new structural model. This study provides valuable insights into the EGF receptor/HER/ErbB kinase structure and interactions, which can guide the design of future therapies.
authors.affiliation: From the Division of Oncology, Department of Medicine, and.
authors.forename: Timothy S
authors.initials: TS
authors.lastname: Collier
authors.affiliation: the Department of Biomedical Engineering and Center for Computational Medicine and Bioinformatics, University of Michigan, Ann Arbor, Michigan 48109.
authors.forename: Karthikeyan
authors.initials: K
authors.lastname: Diraviyam
authors.affiliation: From the Division of Oncology, Department of Medicine, and.
authors.forename: John
authors.initials: J
authors.lastname: Monsey
authors.affiliation: From the Division of Oncology, Department of Medicine, and.
authors.forename: Wei
authors.initials: W
authors.lastname: Shen
authors.affiliation: the Department of Biomedical Engineering and Center for Computational Medicine and Bioinformatics, University of Michigan, Ann Arbor, Michigan 48109.
authors.forename: David
authors.initials: D
authors.lastname: Sept
authors.affiliation: From the Division of Oncology, Department of Medicine, and; the Department of Cell Biology and Physiology, Washington University School of Medicine, St. Louis, Missouri 63110 and. Electronic address: rbose@dom.wustl.edu.
authors.forename: Ron
authors.initials: R
authors.lastname: Bose
mesh_terms: D000818:Animals; D006801:Humans; D013058:Mass Spectrometry; D056004:Molecular Dynamics Simulation; D055503:Protein Multimerization; D020836:Protein Structure, Quaternary; D017433:Protein Structure, Secondary; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D020893:Receptor, ErbB-3 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D061987:Sf9 Cells; D018411:Spodoptera
chemical_list: D018719:Erb-b2 Receptor Tyrosine Kinases; D020893:Receptor, ErbB-3; C508053:ERBB2 protein, human; C581292:ERBB3 protein, human
keywords: Epidermal Growth Factor Receptor (EGFR); Mass Spectrometry (MS); Molecular Dynamics; Protein-Protein Interactions; Receptor Tyrosine Kinase