Out-of-Equilibrium Biophysical Chemistry: The Case for Multidimensional, Integrated Single-Molecule Approaches

Abstract

Out-of-equilibrium processes are ubiquitous across living organisms and all structural hierarchies of life. At the molecular scale, out-of-equilibrium processes (for example, enzyme catalysis, gene regulation, and motor protein functions) cause biological macromolecules to sample an ensemble of conformations over a wide range of time scales. Quantifying and conceptualizing the structure–dynamics to function relationship is challenging because continuously evolving multidimensional energy landscapes are necessary to describe nonequilibrium biological processes in biological macromolecules. In this perspective, we explore the challenges associated with state-of-the-art experimental techniques to understanding biological macromolecular function. We argue that it is time to revisit how we probe and model functional out-of-equilibrium biomolecular dynamics. We suggest that developing integrated single-molecule multiparametric force–fluorescence instruments and using advanced molecular dynamics simulations to study out-of-equilibrium biomolecules will provide a path towards understanding the principles of and mechanisms behind the structure–dynamics to function paradigm in biological macromolecules.

Learning Continuous Potentials from smFRET

“…Also, our method deals with dynamics along a single reaction coordinate assumed to be equivalent to the FRET pair distance. However, one can imagine situations in which the system's dynamics are probed along an axis partly orthogonal to the FRET pair distance (40,41) in a multi-dimensional incarnation of FRET with, say, one donor and multiple acceptor labels. For Manuscript submitted to Biophysical Journal 11 example, even in the case of ACTR binding to NCBD, analyzed in this manuscript, the ACTR may rotate with respect to the NCBD during binding.…”

“…For Manuscript submitted to Biophysical Journal 11 example, even in the case of ACTR binding to NCBD, analyzed in this manuscript, the ACTR may rotate with respect to the NCBD during binding. Cases with multiple degrees of freedom are traditionally studied using multicolor smFRET (41)(42)(43)(44) or by pairing data analysis with molecular dynamics simulations (41,45). In principle, one could use SKIPPER-FRET to infer potentials along degrees of freedom orthogonal to the FRET distance by including some mapping from the desired degree of freedom to the FRET pair distance in equation (8).…”

Challenges in Inferring the Directionality of Active Molecular Processes from Single-Molecule Fluorescence Resonance Energy Transfer Trajectories

“…A solution, on the experimental side, is a development of multidimensional techniques. ,− On the data analysis side, irreversibility is often encoded in non-Markov effects. Such effects, fundamentally, cannot be neglected, and they can be detected with appropriate data analysis. , If discovering and quantifying the directionality of the observed dynamics are the objectives, it may also be desirable to estimate entropy production (eq ) directly from raw experimental time series (e.g., the photon sequence in Figure ) rather than after postprocessing the data using, e.g., hidden Markov models.…”

Cell and Tissue Nanomechanics: From Early Development to Carcinogenesis

“…Depending on the shape and type of DNA 3D structures—the rate and the type of compactization—the DNA molecule may absorb the mechanical energy, which pulls the fiber and unfolds it without the breakdown of the chain [ 70 ]. The chromatin structure also includes specific linker DNA that connects the nucleosomes and potentially play a major role in DNA assembly [ 71 ], and may also absorb various mechanical torsion impacts [ 72 ]. Structural chromatin proteins condense DNA molecules and stabilize its shape and structure [ 72 ]; thus, the enriched regions may have other nuclear mechanics, and the mechanical stress may rearrange the DNA and change its transcription profile [ 73 ].…”

“…The impact of the ECM Young’s modulus is also important for naive mesenchymal stem cells, which can undergo neurogenesis, myogenesis, or osteogenesis under conditions of 1, 10, or 100 kPa, respectively. These are Young’s modulus levels typical for brain tissues, muscle, and bones, respectively (see Figure 3 ) [ 71 ]. Differentiation of skeletal muscle requires cell–ECM interactions [ 88 ].…”