Single-molecule spectroscopy of biological polyelectrolytes

Single-molecule fluorescence spectroscopy has evolved into a versatile method for probing distances, distance distributions, dynamics, and interactions of biopolymers such as proteins and nucleic acids. The chemical tools available for biopolymers facilitate their site-specific labeling with fluorophores, allowing the use of Förster resonance energy transfer (FRET) as a spectroscopic ruler in the nanometer range. I will present the basic concepts of single-molecule spectroscopy and FRET, and then show how these methods can be used to obtain information about biopolymers, especially polyelectrolytes, such as highly charged intrinsically disordered proteins and nucleic acids. In particular, I will show how the combination of such measurements with concepts from polymer physics allows us to quantify a number of interesting properties and processes, including (i) intrachain distance distributions and translational diffusion coefficients, which can be related to radii of gyration and hydrodynamic radii, respectively; (ii) intrachain distance dynamics, which can be related to chain relaxation times of the polymer and the phenomenon of internal friction; (iii) the formation of intermolecular complexes between polyelectrolytes, including the quantification of binding equilibria, kinetics, and the role of counterion release; (iv) the formation of complex coacervates and the conformational properties of biological polyelectrolytes within biomolecular condensates.

References:

Schuler, B., Hofmann, H., Nettels, D. & Soranno, A. (2016) Single-molecule FRET spectroscopy and the polymer physics of unfolded and intrinsically disordered proteins. Annu. Rev. Biophys. 45, 207-231.

Chowdhury, A., Nettels, D. & Schuler, B. (2023) Interaction dynamics of intrinsically disordered proteins from single-molecule spectroscopy. Annu. Rev. Biophys. 52, 433-462.

Nettels, D., Galvanetto, N., Ivanović, M.T., Nüesch, M., Yang, T. & Schuler, B. (2024) Single-molecule FRET for probing nanoscale biomolecular dynamics. Nat. Phys. Rev. 6, 587–605.