Individual Macromolecule Motion in a Crowded Living Cell

The application of the general model of the law of mass action itself, of the achievement of dynamic, statistical equilibria, has led to great successes in describing the theory of single-molecule biophysics and biochemistry based on individually and freely diffusing molecules in dilute liquid and crowded living cells. For example, anomalous diffusion is a general phenomenon in living cells. There is solid evidence for analyzing fluorescence correlation and dual color fluorescence crosscorrelation spectroscopy data ( FCS and dual color FCCS) in cellular applications by equations based on anomalous subdiffusion. Using equations based on normal diffusion causes artifacts of the fitted biological system response parameters and of the interpretations of the FCS and dual color FCCS data in the crowded environment of living cells. Equations based on normal diffusion are not valid in living cells. The original article embraces the status of the experimental situation and touches obstacles that still hinder the applications of single molecules in the cellular environment.

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Foldes-Papp Z

Individual Macromolecule Motion in a Crowded Living Cell

Curr Pharm Biotechnol. 16(1):1-2 (2015). doi: 10.2174/1389201016666141229103953

Anomalous translational diffusion and subdiffusion, respectively, is a breakdown of the laws of mass action. As opposed to normal translational diffusion, in which the movement of molecules is not correlated with their previous position, anomalous translational diffusion molecules are spatially and temporally correlated.

This spatio-temporal correlation reflects a fundamentally different behavior compared with the one in dilute or very dilute solution, which, for example, affects the spread of molecules within live cells.

Until now, characterization of translational diffusion in live cells has relied almost exclusively on measurements of constant translational diffusion coefficients. There is some evidence that in live cells the apparent translational diffusion coefficients may not be constant, but instead can vary over time, even for inert molecules.

In order to take account of temporal randomness of molecular interaction, i.e. time (rate)-dependent sources of anomalous translational diffusion behavior, we decoupled spatial and temporal coordinates of anomalous diffusion. The exponent α (alpha) now quantifies the crowding conditions (spatial heterogeneity) and the exponent γ (gamma) the temporal heterogeneity in anomalous diffusion.

Knowing the crowding parameter α (alpha) for the cell type as well as the cellular compartment, the heterogeneous parameter γ (gamma) can be extracted from the measurements in the presence of the interacting reaction partner, e.g. ligand, for the same (fixed) α value.

Fluorescence correlation spectroscopy ( FCS ) and two-color fluorescence cross-correlation spectroscopy (FCCS) are the methods of choice for measuring the exponents of anomalous subdiffusive translational motion of molecules in live cells as demonstrated for the first time.

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