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An SH3-binding allosteric modulator stabilizes the global conformation of the AML-associated Src-family kinase, Hck.

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Τίτλος: An SH3-binding allosteric modulator stabilizes the global conformation of the AML-associated Src-family kinase, Hck.
Συγγραφείς: Selzer AM; Department of Microbiology and Molecular Genetics, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania, USA., Gerlach G; Department of Computational and Systems Biology, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania, USA., Gonzalez-Areizaga G; Department of Microbiology and Molecular Genetics, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania, USA., Wales TE; Department of Chemistry and Chemical Biology, College of Science, Northeastern University, Boston, Massachusetts, USA., Cui SY; Department of Pharmaceutical Sciences, University of Pittsburgh School of Pharmacy, Pittsburgh, Pennsylvania, USA., Iyer P; Department of Pharmaceutical Sciences, University of Pittsburgh School of Pharmacy, Pittsburgh, Pennsylvania, USA., Engen JR; Department of Chemistry and Chemical Biology, College of Science, Northeastern University, Boston, Massachusetts, USA., Camacho C; Department of Computational and Systems Biology, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania, USA., Ishima R; Department of Structural Biology, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania., Smithgall TE; Department of Microbiology and Molecular Genetics, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania, USA. Electronic address: tsmithga@pitt.edu.
Πηγή: The Journal of biological chemistry [J Biol Chem] 2025 Jan; Vol. 301 (1), pp. 108088. Date of Electronic Publication: 2024 Dec 13.
Τύπος έκδοσης: Journal Article; Research Support, N.I.H., Extramural
Γλώσσα: English
Στοιχεία περιοδικού: Publisher: Elsevier Inc. on behalf of American Society for Biochemistry and Molecular Biology Country of Publication: United States NLM ID: 2985121R Publication Model: Print-Electronic Cited Medium: Internet ISSN: 1083-351X (Electronic) Linking ISSN: 00219258 NLM ISO Abbreviation: J Biol Chem Subsets: MEDLINE
Imprint Name(s): Publication: 2021- : [New York, NY] : Elsevier Inc. on behalf of American Society for Biochemistry and Molecular Biology
Original Publication: Baltimore, MD : American Society for Biochemistry and Molecular Biology
Ιατρικοί όροι (MeSH): Proto-Oncogene Proteins c-hck*/chemistry , Proto-Oncogene Proteins c-hck*/metabolism , Proto-Oncogene Proteins c-hck*/genetics , Proto-Oncogene Proteins c-hck*/antagonists & inhibitors , Leukemia, Myeloid, Acute*/enzymology , Leukemia, Myeloid, Acute*/drug therapy , Leukemia, Myeloid, Acute*/genetics , Protein Kinase Inhibitors*/pharmacology , Protein Kinase Inhibitors*/chemistry , Pyrimidines*/pharmacology , Pyrimidines*/chemistry , src Homology Domains*, Allosteric Regulation/drug effects ; Humans ; Protein Conformation ; Molecular Dynamics Simulation ; Binding Sites
Περίληψη: While ATP-site inhibitors for protein-tyrosine kinases are often effective drugs, their clinical utility can be limited by off-target activity and acquired resistance mutations due to the conserved nature of the ATP-binding site. However, combining ATP-site and allosteric kinase inhibitors can overcome these shortcomings in a double-drugging framework. Here we explored the allosteric effects of two pyrimidine diamines, PDA1 and PDA2, on the conformational dynamics and activity of the Src-family tyrosine kinase Hck, a promising drug target for acute myeloid leukemia. Using 1H-15N HSQC NMR, we mapped the binding site for both analogs to the SH3 domain. Despite the shared binding site, PDA1 and PDA2 had opposing effects on near-full-length Hck dynamics by hydrogen-deuterium exchange mass spectrometry, with PDA1 stabilizing and PDA2 disrupting the overall kinase conformation. Kinase activity assays were consistent with these observations, with PDA2 enhancing kinase activity while PDA1 was without effect. Molecular dynamics simulations predicted selective bridging of the kinase domain N-lobe and SH3 domain by PDA1, a mechanism of allosteric stabilization supported by site-directed mutagenesis of N-lobe contact sites. Cellular thermal shift assays confirmed SH3 domain-dependent interaction of PDA1 with WT Hck in myeloid leukemia cells and with a kinase domain gatekeeper mutant (T338M). These results identify PDA1 as a starting point for Src-family kinase allosteric inhibitor development that may work in concert with ATP-site inhibitors to suppress the evolution of resistance.
(Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)
Competing Interests: Conflict of interest The authors declare that they have no conflicts of interest with the contents of this article.
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Grant Information: F31 CA265294 United States CA NCI NIH HHS; R01 CA233576 United States CA NCI NIH HHS
Contributed Indexing: Keywords: NMR; SH2 domain; SH3 domain; Src-family kinase; hydrogen-deuterium exchange mass spectrometry; kinase inhibitors; tyrosine kinases
Substance Nomenclature: EC 2.7.10.2 (Proto-Oncogene Proteins c-hck)
EC 2.7.10.2 (HCK protein, human)
0 (Protein Kinase Inhibitors)
0 (Pyrimidines)
Entry Date(s): Date Created: 20241215 Date Completed: 20250427 Latest Revision: 20250604
Update Code: 20260130
PubMed Central ID: PMC11786751
DOI: 10.1016/j.jbc.2024.108088
PMID: 39675702
Βάση Δεδομένων: MEDLINE
Περιγραφή
ISSN:1083-351X
DOI:10.1016/j.jbc.2024.108088