DNA Polymerase as a Molecular Motor and Pump

Abstract

DNA polymerase is responsible for synthesizing DNA, a key component in the running of biological machinery. Using fluorescence correlation spectroscopy, we demonstrate that the diffusive movement of a molecular complex of DNA template and DNA polymerase enhances during nucleotide incorporation into the growing DNA template. The diffusion coefficient of the complex also shows a strong dependence on its inorganic cofactor, Mg2+ ions. When exposed to gradients of either nucleotide or cofactor concentrations, an ensemble of DNA polymerase complex molecules shows collective movement toward regions of higher concentrations. By immobilizing the molecular complex on a patterned gold surface, we demonstrate the fabrication of DNA polymerase-powered fluid pumps. These miniature pumps are capable of transporting fluid and tracer particles in a directional manner with the pumping speed increasing in the presence of the cofactor. The role of DNA polymerase as a micropump opens up avenues for designing miniature fluid pumps using enzymes as engines.

Quantifying the Heat Dissipation from Molecular Motor’s Transport Properties in Nonequilibrium Steady States

“…Compared with freely diffusing exothermic enzymes, kinesin-1 whose dynamics is confined on one-dimensional tracks is highly efficient in transforming conformational fluctuations into a locally directed motion, thus displaying a significantly higher enhancement in diffusivity with its turnover rate. Putting molecular motors and freely diffusing enzymes on an equal footing, our study offers thermodynamic basis to understand the heat enhanced self-diffusion of exothermic enzymes.Together with recent studies [1][2][3][4], Riedel et al [5] have demonstrated a rather surprising result, from a perspective of equilibrium statistical mechanics: Diffusion constants (D) of exothermic enzymes, measured from fluorescence correlation spectroscopy (FCS), increase linearly with their catalytic turnover rates V cat , so that the enhancement of diffusivity at maximal activity (maximum V cat ) is as large aswhere D 0 and D max are the diffusion constants measured by FCS at V cat = 0 and maximal V cat , respectively. Their enigmatic observation [5] has called much attention of biophysics community to the physical origin of the activity-dependent diffusivity of a single enzyme.…”

“…Together with recent studies [1][2][3][4], Riedel et al [5] have demonstrated a rather surprising result, from a perspective of equilibrium statistical mechanics: Diffusion constants (D) of exothermic enzymes, measured from fluorescence correlation spectroscopy (FCS), increase linearly with their catalytic turnover rates V cat , so that the enhancement of diffusivity at maximal activity (maximum V cat ) is as large as ∆D/D 0 [≡ (D max − D 0 )/D 0 ] ≈ 0.3 − 3, where D 0 and D max are the diffusion constants measured by FCS at V cat = 0 and maximal V cat , respectively. Their enigmatic observation [5] has called much attention of biophysics community to the physical origin of the activity-dependent diffusivity of a single enzyme.…”

“…Together with recent studies [1][2][3][4], Riedel et al [5] have demonstrated a rather surprising result, from a perspective of equilibrium statistical mechanics: Diffusion constants (D) of exothermic enzymes, measured from fluorescence correlation spectroscopy (FCS), increase linearly with their catalytic turnover rates V cat , so that the enhancement of diffusivity at maximal activity (maximum V cat ) is as large as…”

Quantitative Analysis of Purine Nucleotides Indicates That Purinosomes Increase de Novo Purine Biosynthesis

“…Two possibilities remain to be explored. The first is reversible posttranslational modification of one or more member of these enzymes; the second is substrate concentration-dependent increase in diffusion of the enzymes in the presence of their respective substrates to drive complex formation (Brownian ratchet) (35)(36)(37). Both are being studied.…”

Tight Coupling between the Heat Dissipation and Molecular Motor's Transport Properties in Nonequilibrium Steady State

“…The philosophy underlying the analysis of mapping trajectories on kinetic model, proposed here on kinesin-1 as well as others on F 1 -ATPase [29,46], is in essence similar to the one by the recent study which has quantified circulating flux on configurational phase space (or mode space) to diagnose broken DB and non-equilibrium dynamics 4 0.14 at mesoscopic scale [37,38]. Lastly, our study confers quantitative insights into how much of the chemical free energy supplied to active systems (enzymes, molecular motors) is converted to mechanical movement in space and eventually dissipated into heat.…”