DOI: 10.1101/483321Nov 30, 2018Paper

Resolving Cytosolic Diffusive States in Bacteria by Single-Molecule Tracking

BioRxiv : the Preprint Server for Biology
Julian M RochaAndreas Gahlmann

Abstract

The trajectory of a single protein in the cytosol of a living cell contains information about its molecular interactions in its native environment. However, it has remained challenging to accurately resolve and characterize the diffusive states that can manifest in the cytosol using analytical approaches based on simplifying assumptions. Here, we show that multiple intracellular diffusive states can be successfully resolved if sufficient single-molecule trajectory information is available to generate well-sampled distributions of experimental measurements and if experimental biases are taken into account during data analysis. To address the inherent experimental biases in camera-based and MINFLUX-based single-molecule tracking, we use an empirical data analysis framework based on Monte Carlo simulations of confined Brownian motion. This framework is general and adaptable to arbitrary cell geometries and data acquisition parameters employed in 2D or 3D single-molecule tracking. We show that, in addition to determining the diffusion coefficients and populations of prevalent diffusive states, the timescales of diffusive state switching can be determined by stepwise increasing the time window of averaging over subsequent single-mol...Continue Reading

Related Concepts

Environment
Intracellular
Cytoplasmic antibody
Three-dimensional
Tracking
Diffusion Weighted Imaging
Simulation
Analysis
Binding (Molecular Function)
Small Molecule

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