bioRxiv Science⌕ Search

Biology subjects

Muthukumar, M.

Publications and source records attributed to Muthukumar, M..

8 recordsLinked to original sources

Topologically frustrated dynamics in an uncharged Tetra-PEG gel

Based on our previous discovery of non-diffusive topologically frustrated dynamics in a charged system where electrostatic interactions between the charged guest and the charged host gel play a role (Nature Communications, 2018, 9, 2248; Physical Review Letters, 2021, 126, 057802), we have investigated the onset of this effect in an uncharged gel matrix. Using sodium (polystyrene sulfonate) as the guest macromolecule and the ideal tetra-PEG gel, we find the emergence of the non-diffusive topologically frustrated dynamical state with a hierarchy of segmental dynamics represented by a stretched exponential of exponent {beta} around 1/3. Our results demonstrate the universal behavior of the topologically frustrated dynamical state.

biophysics↗

Nucleotide Dynamics During Flossing of Polycation-DNA-Polycation through a Nanopore using Molecular Dynamics

The full potential of solid state nanopores is yet to be realized for genome sequencing. Due to its robustness it can handle strong voltage amplitude and frequency. The effect of strong alternating voltage on the dynamics of nucleotides during translocation has been explored. We proposed a setup consisting of single stranded DNA covalently linked with symmetric polycations at both ends fashioned after the proposal of Kasianowicz. 1 Such a setup allows for repeated back and forth motion of the DNA along the nanopore (1.45 nm diameter and 1.53 nm thick) by simply switching the voltage polarity if the polycation tail is sufficiently long ([≥] 10) and the applied voltage is below 0.72 volts, but the average residence time of the nucleotides are too small to be of any practical use (6-30 ns). When alternating voltage of higher frequency is applied, it enhances the average residence time of the nucleotides by an order of magnitude to [~] 0.1 {micro}s relative to direct voltage but the individual trajectories are too stochastic. Since, we are able to collect repeated read on the dynamics of individual nucleotides, we obtained the most probable time of appearance of a nucleotide within the nanopore. With such construct we were able to get almost linear dependence of most probable time versus nucleotide index, after gaussian fitting.

biophysics↗

Langevin dynamics simulation of protein dynamics in nanopores at microsecond timescales

With rapid advancement in the fields of nanopore analysis of protein, it has become imperative to develop modeling framework for understanding the protein dynamics in nanopores. Such modeling framework should include the effects of electro-osmosis, as it plays significant role during protein translocation in confinement. Currently, the molecular dynamics simulations that include the hydrodynamic effects are limited to a timescale of few 100 ns. These simulations give insight about important events like protein unfolding which occurs in this timescale. But many electrophoresis experiments are limited by a detector resolution of ~ 2.5 s. Analytical theory has been used to interpret protein dynamics at such large timescale. There is a need for molecular modeling of more complex environment and protein shapes which cannot be accounted for by analytical theory. We have developed a framework to study globular protein dynamics in nanopores by using langevin dynamics on a rigid body model of protein and the hydrodynamics is accounted by analytical theory for simple cylindrical nanopore geometry. This framework has been applied to study the dynamics of Ubiquitin translocation in SiNx nanopore by Nir et al26. They have reported 7 times decrease in average dwell time of the protein inside the nanopore in response to a small change in pH from 7.0 to 7.2 and the modification of protein charge was attributed for such drastic change. Closer examination using our simulation revealed that the electro-osmotic effects originating due to very small change in the surface electrostatic potential of the nanopore could lead to such a drastic change in protein dynamics.

biophysics↗

Coupled dynamics of charged macromolecules and counterions mediated by binding enzymes

We investigate the role of active coupling on the transport properties of the macromolecules. The active coupling comes due to bound enzymes with a segment of the macromolecule wherein the enzyme exerts an electrostatic force on the segment of the macromolecule, and eventually, it gets unbound due to the thermal fluctuations. This binding and unbinding process generates active fluctuations in the dynamics of the macromolecule. Starting with segment dynamics and correlations for three dynamical models with active coupling, we obtain the cooperative diffusivity for the realistic charged macromolecules with hydrodynamics. First, we construct the three models by incorporating the features of a real polymer systematically, starting from simple Rouse dynamics with active coupling. We further include segment-segment interactions and in addition, hydrodynamic interactions with active coupling. Our obtained scaling form for segment-segment correlations for the models in terms of the size exponent of the polymer indicating that hydrodynamic and segment-segment interactions along with the active coupling lead to new scaling regimes. We finally study the dynamics of a homogeneously charged flexible polymer in an infinitely dilute solution where enzymes and counterions affect the dynamics of the polymers. We analytically investigate how these active fluctuations affect the coupled dynamics of the polymer and counterions. It turns out that these active fluctuations enhance the effective diffusivity of the polymer. The derived closed-form expression for diffusivity is pertinent to accurate interpretation of light scattering data on multi-component systems with binding-unbinding equilibria.

biophysics↗

Voltage-driven polyelectrolyte complexation inside a nanopore

We have investigated how a pair of oppositely charged macromolecules can be driven by an electric field to form a polyelectrolyte complex inside a nanopore. To observe and isolate an individual complex pair, a model protein nanopore, embedded in artificial phospholipid membrane, allowing compartmentalization (cis/trans) is employed. A polyanion in the cis and a polycation in the trans compartments are subjected to electrophoretic capture by the pore. We find that the measured ionic current across the pore has a distinguishable signature of complex formation, which is different from the signature of the passage of individual molecules through the pore. The ionic current signature allows us to detect the interaction between the two oppositely charged macromolecules and thus, enables us to measure the lifetime of the complex inside the nanopore. After showing that we can isolate a complex pair in the nanopore, we studied the effects of molecular identity on the nature of interaction in different complex pairs. In contrast to the irreversible conductance state of the alpha-hemolysin (HL) channel in the complexation of poly-styrene-sulfonate (PSS) and poly-L-lysine (PLL), a reversible conductance state is observed during complexation between single stranded DNA (ssDNA) and PLL. This suggests that there is a weak interaction between ssDNA and PLL, when compared to the interaction in a PSS-PLL complex. Analysis of the PSS-PLL complexation events and its lifetime inside the nanopore supports a four step-mechanism: (i) The polyanion is captured by the pore, (ii) the polyanion starts threading through the pore. (iii) The polycation is captured, a complex pair is formed in the pore, and the polyanion slides along the polycation. (iv) The complex pair can be pulled through the pore into the trans compartment or it can dissociate. Additionally, we have developed a simple theoretical model, which describes the lifetime of the complex inside the pore. The observed reversible two-state conductance across HL channel during ssDNA-PLL complexation, is described as the binding/unbinding of PLL during the translocation of ssDNA. This enables us to evaluate the apparent rate constants for association/dissociation and equilibrium dissociation constants for the interaction of PLL with ssDNA. This work throws light on the behavior of polyelectrolyte complexes in an electric field and enhances our understanding of the electrical aspects of inter-macromolecular interactions, which plays an extremely important role in the organization of macromolecules in the crowded and confined cellular environment.

biophysics↗

Stochastic Resonance Behavior of DNA Translocation with an Oscillatory Electric Field

Stochastic resonance (SR) describes the synchronization between noise of a system and an applied oscillating field to achieve an optimized response signal. In this work, we use simulations to investigate the phenomenon of SR of a single stranded DNA driven through a nanopore when an oscillating electric field is added. The system is comprised of a MspA protein nanopore embedded in a membrane and different lengths of DNA is driven from one end of the pore to the other via a constant potential difference. We superimposed an oscillating electric field on top of the existing electric field. The source of noise is due to thermal fluctuations, since the system is immersed in solution at room temperature. Here, the signal optimization we seek is the increase in translocation time of DNA through the protein nanopore. Normally, translocation time scales linearly with DNA length and inversely with driving force in a drift dominated regime. We found a non-monotonic dependence of the mean translocation time with the frequency of the oscillating field. This non-monotonic behavior of the translocation time is observed for all lengths of DNA, but SR occurs only for longer DNA. Furthermore, we also see evidence of DNA extension being influenced by the oscillating field while moving through the nanopore.

biophysics↗

Unfolding of RNA via Translocation Through a Nanopore

RNA unfolding and refolding are important biological phenomena, which occur during the transfer of genetic information from DNA to RNA to proteins. During these processes, RNA is found in single stranded, secondary and tertiary structures, including secondary conformations like hairpins and pseudoknots. Understanding the diverse conformations of RNA and how these influence the dynamics of unfolding and refolding is crucial to gain insight to fundamental biological processes. In this work, we employ coarse-grained Langevin dynamics simulations of a simple model of different RNA hairpins passing through a geometric nanopore to find the influence of structural changes on the translocation dynamics. The threshold voltage of unfolding depends on the length of the hairpin attached to the tail. The lag time to unfold is longer for smaller applied voltages and for the architectures containing a longer hairpin attached to the tail. Chain translocation dynamics for different architectures are largely collapsed by the threshold. A distinct signature for the base unfolding time was observed for the bases around the unpaired bases in all the RNA hairpin models. These results can motivate future technologies or experiments that use translocation to predict secondary structures of polynucleotides.

biophysics↗

Macromolecule Translocation in a Nanopore: Center of Mass Drift-Diffusion over an Entropic Barrier

Many fundamental biological processes involve moving macromolecules across membranes, through nanopores, in a process called translocation. Such motion is necessary for gene expression and regulation, tissue formation, and viral infection. Furthermore, in recent years nanopore technologies have been developed for single molecule detection of biological and synthetic macromolecules, which have been most notably employed in next generation DNA sequencing devices. Many successful theories have been established, which calculate the entropic barrier required to elongate a chain during translocation. However, these theories are at the level of the translocation coordinate (number of forward steps) and thus lack a clear connection to experiments and simulations. Furthermore, the proper diffusion coefficient for such a coordinate is unclear. In order to address these issues, we propose a center of mass (CM) theory for translocation. We start with the entropic barrier approach and show that the translocation coordinate is equivalent to the center of mass of the chain, providing a direct interpretation of previous theoretical studies. We thus recognize that the appropriate dynamics is given by CM diffusion, and calculate the appropriate diffusion constant (Rouse or Zimm) as the chain translocates. We illustrate our theoretical approach with a planar nanopore geometry and calculate some characteristic dynamical predictions. Our main result is the connection between the translocation coordinate and the chain CM, however, we also find that the translocation time is sped up by 1-2 orders of magnitude if hydrodynamic interactions are present. Our approach can be extended to include the details included in previous translocation theories. Most importantly this work provides a direct connection between theoretical approaches and experiments or simulations.\n\nSIGNIFICANCEMacromolecule motion through nanopores is critical for many biological processes, and has been recently employed for nucleic acid sequencing. Despite this, direct theoretical understandings of translocation are difficult to evaluate due to the introduction of the translocation coordinate. In this manuscript, we propose a theory for translocation written at the center of mass level of the polymer chain. This theoretical approach is more easily compared to experimental and simulation results, and additionally allows one to accurately account for hydrodynamic interactions on the macromolecule dynamics.

biophysics↗