Biology subjects

Prakaash, D.

Publications and source records attributed to Prakaash, D..

3 recordsLinked to original sources

bioRxiv2022

Molecular dynamics simulations reveal membrane lipid interactions of the full-length lymphocyte specific kinase Lck

The membrane-bound lymphocyte-specific protein-tyrosine kinase (Lck) triggers T cell antigen receptor signalling to initiate adaptive immune responses. Despite many structure-function studies, the mode of action of Lck and the potential role of plasma membrane lipids in regulating Lcks activity remains elusive. Advances in molecular dynamics simulations of membrane proteins in complex lipid bilayers have opened a new perspective in gathering such information. Here, we have modelled the full-length Lck open and closed conformations available from crystallographic studies and simulated its interaction with the inner leaflet of the T cell plasma membrane. In both conformations, we found that the unstructured unique domain and the structured domains including the kinase interacted with the membrane with a preference for PIP lipids. Interestingly, our simulations suggest that the Lck-SH2 domain interacts with lipids differently in the open and closed Lck conformations, demonstrating that lipid interaction can potentially regulate Lcks conformation and in turn modulate T cell signalling. Additionally, the Lck-SH2 and kinase domain residues that significantly contacted PIP lipids are found to be conserved among the Src family of kinases, thereby potentially representing similar PIP interactions within the family.

biophysics↗

bioRxiv2021

Multi-scale simulations of the T cell receptor reveal its lipid interactions, dynamics and the arrangement of its cytoplasmic region

The T cell antigen receptor (TCR-CD3) complex initiates T cell activation following recognition of peptides presented by Major Histocompatibility Complex (pMHC)-encoded proteins. The ligation of pMHC to TCR{beta} induces Src family kinases activity via the cytoplasmic tails of the CD3{delta}{varepsilon}, CD3{gamma}{varepsilon} and {zeta}{zeta} dimers. The TCR-CD3 topology is well understood, but little is known about its conformational dynamics and arrangement of its cytoplasmic tails, limiting our grasp of the signalling mechanism. Here, we investigated the entire TCR-CD3 embedded in an asymmetric lipid bilayer using molecular modelling and multi-scale molecular dynamics simulations. Our study demonstrates conformational changes in the extracellular and transmembrane domains, and the arrangement of the TCR-CD3 cytoplasmic tails. The TCR{beta} variable regions were the most flexible in the extracellular domain. The cytoplasmic tails formed highly interlaced structures while some tyrosine sidechains within the immunoreceptor tyrosine-based activation motifs (ITAMs) of the CD3{varepsilon} and {zeta} subunits dynamically penetrated the hydrophobic core of the bilayer. Ionic interactions between the cytoplasmic tails and phosphatidylinositol phosphates (PIP2 and PIP3) in the inner leaflet of the lipid bilayer led to the formation of a distinct annular lipid fingerprint around the TCR-CD3 complex. These results combined with available experiential data increase our understanding of the TCR-CD3 activation mechanism and highlight the importance of membrane lipids in regulating T cell activation. Significance statementThe T cell receptor (TCR-CD3) detects antigenic peptides displayed by major histocompatibility complexes (pMHC) to instigate activation of T cell adaptive immunity. Despite significant structural and functional knowledge of TCR-CD3 topology, the membrane interactions and dynamics of its cytoplasmic moieties remain elusive. Interactions of TCR-CD3 cytoplasmic tails with membrane lipids may regulate their phosphorylation by Src-family kinases, the first intracellular event required for T cell activation. Using the static 3D structure of TCR-CD3 resolved by cryo-electron microscopy, we provide novel insights into the protein-lipid interactions of the complete TCR-CD3 embedded in a bilayer closely mimicking its native membrane environment. Our study sheds light on the dynamics of the TCR-CD3 at near-atomic resolution and further aids in deciphering its activation mechanism.

biophysics↗

bioRxiv2020

Allosteric activation of T cell antigen receptor signalling by quaternary structure relaxation

The mechanism of T cell antigen receptor (TCR-CD3) signalling remains elusive. Here, we identified mutations in the transmembrane region of TCR{beta} or CD3{zeta} that augmented pMHC-induced signalling, not explicable by enhanced ligand binding, lateral diffusion, clustering or co-receptor function. Using a novel biochemical assay and molecular dynamics simulation, we demonstrated that the gain-of-function mutations loosened interaction between TCR{beta} and CD3{zeta}. We found that, similar to the activating mutations, pMHC binding reduced TCR{beta} cohesion with CD3{zeta}. This event occurred prior to CD3{zeta} phosphorylation and at 0{degrees}C. Moreover, we demonstrated that soluble monovalent pMHC alone induced signalling and reduced TCR{beta} cohesion with CD3{zeta} in membrane-bound or solubilised TCR-CD3. Our data provide compelling evidence that pMHC binding suffices to activate allosteric changes propagating from TCR{beta} to the CD3 subunits, reconfiguring interchain transmembrane region interactions. These dynamic modifications could change the arrangement of TCR-CD3 boundary lipids to licence CD3{zeta} phosphorylation and initiate signal propagation.

immunology↗