Quantitative Analysis of Protein-Protein Equilibrium Constants in Cellular Environments Using Single-Molecule Localization Microscopy.

ABSTRACT: Current methods for determining equilibrium constants often operate in three-dimensional environments, which may not accurately reflect interactions with membrane-bound proteins. With our technique, based on single-molecule localization microscopy (SMLM), we directly determine protein-protein association (K_a) and dissociation (K_d) constants in cellular environments by quantifying associated and isolated molecules and their interaction area. We introduce Kernel Surface Density (ks-density,) a novel method for determining the accessible area for interacting molecules, eliminating the need for user-defined parameters. Simulation studies validate our method's accuracy across various density and affinity conditions. Applying this technique to T cell signaling

SUBMITTER: Marcano-Garcia LF 

PROVIDER: S-EPMC11528428 | biostudies-literature | 2024 Oct

REPOSITORIES: biostudies-literature