Biology subjects

Chung, J. K.

Publications and source records attributed to Chung, J. K..

4 recordsLinked to original sources

bioRxiv2026

Small-Molecule Inhibitors Allosterically Control Bruton's Tyrosine Kinase Assembly on the Membrane

Bruton's tyrosine kinase (Btk) is the target of small-molecule, active site inhibitors for treatment of lymphomas and leukemias. Beyond its catalytic activity, Btk participates in signaling assemblies through scaffolding functions that can contribute to drug resistance. Here, we show that Btk forms micron-scale assemblies on membranes and that this behavior is allosterically controlled by ligand binding at the active site. Using supported lipid bilayers and total internal reflection fluorescence microscopy, we find that nucleotides and inhibitors including ibrutinib, acalabrutinib, and dasatinib promote assembly, whereas fenebrutinib and pirtobrutinib prevent assembly. Membrane organization is independent of catalysis but correlates with ligand-dependent changes in Btk conformational flexibility measured by hydrogen-deuterium exchange. These results reveal a previously unrecognized connection between active-site ligand binding and mesoscale membrane organization, demonstrating that kinase inhibitors can control Btk behavior beyond suppression of catalytic activity.

biophysics↗

bioRxiv2020

Coupling between lipid miscibility and phosphotyrosine driven protein condensation at the membrane

Lipid miscibility phase separation has long been considered to be a central element of cell membrane organization. More recently, protein condensation phase transitions, into three-dimensional droplets or in two-dimensional lattices on membrane surfaces, have emerged as another important organizational principle within cells. Here, we reconstitute the LAT:Grb2:SOS protein condensation on the surface of giant unilamellar vesicles capable of undergoing lipid phase separations. Our results indicate that assembly of the protein condensate on the membrane surface can drive lipid phase separation. This phase transition occurs isothermally and is governed by tyrosine phosphorylation on LAT. Furthermore, we observe that the induced lipid phase separation drives localization of the SOS substrate, K-Ras, into the LAT:Grb2:SOS protein condensate. Statement of SignificanceProtein condensation phase transitions are emerging as an important organizing principles in cells. One such condensate plays a key role in T cell receptor signaling. Immediately after receptor activation, multivalent phosphorylation of the adaptor protein LAT at the plasma membrane leads to networked assembly of a number of signaling proteins into a two-dimensional condensate on the membrane surface. In this study, we demonstrate that LAT condensates in reconstituted vesicles are sufficient to drive lipid phase separation. This lipid reorganization drives another key downstream signaling molecule, Ras, into the LAT condensates. These results show that the LAT condensation phase transition, which is actively controlled by phosphorylation reactions, extends its influence to control lipid phase separation in the underlying membrane.

biophysics↗

bioRxiv2020

Raf promotes dimerization of the Ras G-domain with increased allosteric connections

Ras dimerization is critical for Raf activation, yet Ras alone does not dimerize. Here we show that the Ras binding domain of Raf (Raf-RBD) induces robust Ras dimerization at low surface densities on supported lipid bilayers and, to a lesser extent, in solution as observed by size exclusion chromatography and confirmed by SAXS. Community network analysis based on molecular dynamics (MD) simulations show robust allosteric connections linking the two Raf-RBD D113 residues, located in the Galectin scaffold protein binding site of each Raf-RBD molecule and 85 [A] apart on opposite ends of the dimer complex. Our results suggest that Raf-RBD binding and Ras dimerization are concerted events that lead to a high-affinity signaling complex at the membrane that we propose is an essential unit in the macromolecular assembly of higher order Ras/Raf/Galectin complexes important for signaling through the Ras/Raf/MEK/ERK pathway.

biophysics↗

bioRxiv2018

Switch-like activation of Bruton’s tyrosine kinase by membrane-mediated dimerization

The transformation of molecular binding events into cellular decisions is the basis of most biological signal transduction. A fundamental challenge faced by these systems is that protein-ligand chemical affinities alone generally result in poor sensitivity to ligand concentration, endangering the system to error. Here, we examine the lipid-binding pleckstrin homology and Tec homology (PH-TH) module of Brutons tyrosine kinase (Btk) Using fluorescence correlation spectroscopy (FCS) and membrane-binding kinetic measurements, we identify a self-contained phosphatidylinositol (3,4,5)-trisphosphate (PIP3) sensing mechanism that achieves switch-like sensitivity to PIP3 levels, surpassing the intrinsic affinity discrimination of PIP3:PH binding. This mechanism employs multiple PIP3 binding as well as dimerization of Btk on the membrane surface. Mutational studies in live cells confirm that this mechanism is critical for activation of Btk in vivo. These results demonstrate how a single protein module can institute a minimalist coincidence detection mechanism to achieve high-precision discrimination of ligand concentration.

biophysics↗