Abstract
c-Src kinase is a multidomain non-receptor tyrosine kinase that aberrantly phosphorylates several signaling proteins in cancers. Although the structural properties of the regulatory domains (SH3-SH2) and the catalytic kinase domain have been extensively characterized, there is less knowledge about the N-terminal disordered region (SH4UD) and its interactions with the other c-Src domains. Here, we used domain-selective isotopic labeling combined with the small-angle neutron scattering contrast matching technique to study SH4UD interactions with SH3-SH2. Our results show that in the presence of SH4UD, the radius of gyration (Rg) of SH3-SH2 increases, indicating that it has a more extended conformation. Hamiltonian replica exchange molecular dynamics simulations provide a detailed molecular description of the structural changes in SH4UD-SH3-SH2 and show that the regulatory loops of SH3 undergo significant conformational changes in the presence of SH4UD, while SH2 remains largely unchanged. Overall, this study highlights how a disordered region can drive a folded region of a multidomain protein to become flexible, which may be important for allosteric interactions with binding partners. This may help in the design of therapeutic interventions that target the regulatory domains of this important family of kinases.
| Original language | English |
|---|---|
| Pages (from-to) | 714-723 |
| Number of pages | 10 |
| Journal | Biomacromolecules |
| Volume | 24 |
| Issue number | 2 |
| DOIs | |
| State | Published - Feb 13 2023 |
Funding
This manuscript has been authored by UT-Battelle, LLC, under contract no. DE-AC05-00OR22725 with the U.S. Department of Energy. The United States Government retains and the publisher, by accepting the article for publication, acknowledges that the United States Government retains a non-exclusive, paid-up, irrevocable, worldwide license to publish or reproduce the published form of this manuscript, or allow others to do so, for United States Government purposes. The Department of Energy will provide public access to these results of federally sponsored research in accordance with the DOE Public Access Plan ( http://energy.gov/downloads/doe-public-access-plan ). Acknowledgments This work was supported by the Laboratory Directed Research and Development Program of Oak Ridge National Laboratory, managed by UT-Battelle LLC, and by project ERKPA14 funded by the Office of Biological & Environmental Research in the Department of Energy (DOE) Office of Science. The authors thank Dr. Kevin Weiss for technical support for bio-deuteration. V.G. acknowledges the support of University of Tennessee Knoxville Science Alliance agency for providing financial support through a Graduate Advancement & Training Education (GATE) fellowship. Neutron scattering experiments on Bio-SANS were supported by the Center for Structural Molecular Biology funded by DOE BER project ERKP291. Computational work was supported by the National Energy Research Scientific Computing Center (contract no. DE-AC02-05CH11231) and The Oak Ridge Leadership Computing Facility (contract no. DE-AC05-00OR22725). This research used resources at the High Flux Isotope Reactor and Spallation Neutron Source, a DOE Office of Science User Facility operated by the Oak Ridge National Laboratory.
| Funders | Funder number |
|---|---|
| DOE BER | ERKP291 |
| Oak Ridge National Laboratory | DE-AC05-00OR22725 |
| University of Tennessee Knoxville Science Alliance | |
| U.S. Department of Energy | |
| Office of Science | |
| Biological and Environmental Research | |
| Oak Ridge National Laboratory | |
| UT-Battelle | |
| National Energy Research Scientific Computing Center | DE-AC02-05CH11231 |
