Luis F. Marcano-García, Cecilia Zaza, Olivia P. L. Dalby, Megan D. Joseph, M. Victoria Cappellari, Sabrina Simoncelli, Pedro F. Aramendía
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引用次数: 0
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 (Ka) and dissociation (Kd) 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 proteins, we determine the 2D association constant of T cell receptors (TCRs) in resting cells and the pseudo-3D dissociation constant of pZAP70 molecules from phosphorylated intracellular tyrosine-based activation motifs on the TCR-CD3 complex. We address challenges of multiple detection and molecular labeling efficiency. This method enhances our understanding of protein interactions in cellular environments, advancing our knowledge of complex biological processes.
期刊介绍:
Nano Letters serves as a dynamic platform for promptly disseminating original results in fundamental, applied, and emerging research across all facets of nanoscience and nanotechnology. A pivotal criterion for inclusion within Nano Letters is the convergence of at least two different areas or disciplines, ensuring a rich interdisciplinary scope. The journal is dedicated to fostering exploration in diverse areas, including:
- Experimental and theoretical findings on physical, chemical, and biological phenomena at the nanoscale
- Synthesis, characterization, and processing of organic, inorganic, polymer, and hybrid nanomaterials through physical, chemical, and biological methodologies
- Modeling and simulation of synthetic, assembly, and interaction processes
- Realization of integrated nanostructures and nano-engineered devices exhibiting advanced performance
- Applications of nanoscale materials in living and environmental systems
Nano Letters is committed to advancing and showcasing groundbreaking research that intersects various domains, fostering innovation and collaboration in the ever-evolving field of nanoscience and nanotechnology.