Robert Hołyst

Comparative analysis of viscosity of complex liquids and cytoplasm of mammalian cells at the nanoscale.

We present a scaling formula for size-dependent viscosity coefficients for proteins, polymers, and fluorescent dyes diffusing in complex liquids. The formula was used to analyze the mobilities of probes of different sizes in HeLa and Swiss 3T3 mammalian cells. This analysis unveils in the cytoplasm two length scales: (i) the correlation length ξ (approximately 5 nm in HeLa and 7 nm in Swiss 3T3 cells) and (ii) the limiting length scale that marks the crossover between nano- and macroscale viscosity (approximately 86 nm in HeLa and 30 nm in Swiss 3T3 cells). During motion, probes smaller than ξ experienced matrix viscosity: η(matrix) ≈ 2.0 mPa·s for HeLa and 0.88 mPa·s for Swiss 3T3 cells. Probes much larger than the limiting length scale experienced macroscopic viscosity, η(macro) ≈ 4.4 × 10(-2) and 2.4 × 10(-2) Pa·s for HeLa and Swiss 3T3 cells, respectively. Our results are persistent for the lengths scales from 0.14 nm to a few hundred nanometers.

Phase behavior of gradient copolymers

Phase properties of gradient AB copolymer melts which consist of chains with the specified chemical distribution of A and B monomers have been studied within the Landau–Ginzburg model. All the melts with the linear distribution of the monomers exhibit only a direct continuous phase transition from disordered to the lamellar phase. The hexagonal, body-centered-cubic, double-gyroid (G), and lamellar-ordered structures have been found in the melts with the monotonic but nonlinear distribution of the monomers. The G structure has been also found in the gradient copolymer melts with the distribution function of monomers similar to the A–B–A triblock copolymers.

Configurational transition in a Fleming - Viot-type model and probabilistic interpretation of Laplacian eigenfunctions

We analyse and simulate a two-dimensional Brownian multi-type particle system with death and branching (birth) depending on the position of particles of different types. The system is confined in a two-dimensional box, whose boundaries act as the sink of Brownian particles. The branching rate matches the death rate so that the total number of particles is kept constant. In the case of m types of particle in a rectangular box of size and elongated shape we observe that the stationary distribution of particles corresponds to the mth Laplacian eigenfunction. For smaller elongations a > b we find a configurational transition to a new limiting distribution. The ratio a/b for which the transition occurs is related to the value of the mth eigenvalue of the Laplacian with rectangular boundaries.

Biologistics--diffusion coefficients for complete proteome of Escherichia coli.

Motivation: Biologistics provides data for quantitative analysis of transport (diffusion) processes and their spatio-temporal correlations in cells. Mobility of proteins is one of the few parameters necessary to describe reaction rates for gene regulation. Although understanding of diffusion-limited biochemical reactions in vivo requires mobility data for the largest possible number of proteins in their native forms, currently, there is no database that would contain the complete information about the diffusion coefficients (DCs) of proteins in a given cell type. Results: We demonstrate a method for the determination of in vivo DCs for any molecule—regardless of its molecular weight, size and structure—in any type of cell. We exemplify the method with the database of in vivo DC for all proteins (4302 records) from the proteome of K12 strain of Escherichia coli, together with examples of DC of amino acids, sugars, RNA and DNA. The database follows from the scale-dependent viscosity reference curve (sdVRC). Construction of sdVRC for prokaryotic or eukaryotic cell requires ~20 in vivo measurements using techniques such as fluorescence correlation spectroscopy (FCS), fluorescence recovery after photobleaching (FRAP), nuclear magnetic resonance (NMR) or particle tracking. The shape of the sdVRC would be different for each organism, but the mathematical form of the curve remains the same. The presented method has a high predictive power, as the measurements of DCs of several inert, properly chosen probes in a single cell type allows to determine the DCs of thousands of proteins. Additionally, obtained mobility data allow quantitative study of biochemical interactions in vivo. Contact: rholyst@ichf.edu.pl Supplementary information: Supplementary data are available at Bioinformatics Online.

Highly reproducible, stable and multiply regenerated surface-enhanced Raman scattering substrate for biomedical applications

We fabricated a Surface Enhanced Raman Scattering (SERS)-active surface based on photo-etched and Au-coated GaN. The highest enhancement factor (EF) in SERS and high reproducibility of spectra were obtained from surfaces covered with bunched nanopillars which were produced by relatively long defect-selective photo-etching. The surfaces exhibited SERS enhancements of the order of 2.8 × 106 for malachite green isothiocyanate (MGITC) and 2 × 106 for p-mercaptobenzoic acid (PMBA). These SERS enhancement factors were comparable to those of conventional SERS substrates, while the EF for MGITC was two orders of magnitude larger than the corresponding one reported for the SERS platform made on porous GaN. The standard deviation of the relative intensity of the 1180 cm−1 mode of MGITC was less than 5% for 100 randomly distributed locations across a single platform and less than 10% between different platforms. The SERS signal of MGITC at our GaN/Au surface (kept under ambient conditions) was extremely stable. We could not detect any peak shift or appreciable change of intensity even after three months. We used these surfaces to detect biological molecules such as amino acids and bovine serum albumin (BSA) at low concentration and with short detection time. We developed simple and effective cleaning procedures for our substrates. After cleaning, the same substrate could be used multiple times retaining the SERS activity. We are not aware of any other multiply regenerated SERS substrate which provides simultaneously such high stability with high enhancement, good uniformity, and high reproducibility.

Study of the Landau bicritical point in dense systems of hard biaxial molecules

We study two systems of hard biaxial molecules: hard spheroplatelets and hard ellipsoids, using the liquid-crystalline version of the smoothed density approximation (SDA). The first system is studied for all elongations of the spheroplatelet c, whereas the second only for c ⩽ 7. For both systems, we locate the line of Landau bicritical points at which a direct transition from the isotropic phase to the biaxial phase occurs. We find that the density of the isotropic phase at the Landau bicritical point is always higher than that at the isotropic-nematic transition in the limit of uniaxial molecules and the difference ranges from 10 to 30 per cent. For hard ellipsoids, we obtain a similar scaling behaviour at the Landau bicritical point as for spheroplatelets, i.e. b ∼c 1/2, where b denotes the breadth of the ellipsoid.

Quantitative influence of macromolecular crowding on gene regulation kinetics

We introduce macromolecular crowding quantitatively into the model for kinetics of gene regulation in Escherichia coli. We analyse and compute the specific-site searching time for 180 known transcription factors (TFs) regulating 1300 operons. The time is between 160 s (e.g. for SoxS Mw = 12.91 kDa) and 1550 s (e.g. for PepA6 of Mw = 329.28 kDa). Diffusion coefficients for one-dimensional sliding are between for large proteins up to for small monomers or dimers. Three-dimensional diffusion coefficients in the cytoplasm are 2 orders of magnitude larger than 1D sliding coefficients, nevertheless the sliding enhances the binding rates of TF to specific sites by 1–2 orders of magnitude. The latter effect is due to ubiquitous non-specific binding. We compare the model to experimental data for LacI repressor and find that non-specific binding of the protein to DNA is activation- and not diffusion-limited. We show that the target location rate by LacI repressor is optimized with respect to microscopic rate constant for association to non-specific sites on DNA. We analyse the effect of oligomerization of TFs and DNA looping effects on searching kinetics. We show that optimal searching strategy depends on TF abundance.

Size and shape of micelles studied by means of SANS, PCS, and FCS.

The hexaethylene glycol monododecyl ether (C(12)E(6)) micelles at concentrations up to 10% have been studied in their isotropic phase (10-48 degrees C) by means of small angle neutron scattering (SANS) and photon correlation spectroscopy (PCS). The SANS data obtained at low temperatures could be unequivocally interpreted as a result of scattering from a suspension of compact globular micelles with the shape of a triaxial ellipsoid or a short end-capped elliptical rod. Different models have been applied to analyze the SANS data obtained at higher temperatures: (i) elongated rod-like micelles with purely sterical interactions, (ii) compact globular micelles with a weak attractive potential, and (iii) globular micelles influenced by the critical phenomena in the whole temperature range studied. The good quality of the experimental data indicated model (i) as the best fit for our data. The diffusion coefficients obtained from the PCS measurements have been compared to the diffusion coefficients calculated for the rod-like micelles--results of the SANS data analysis. A good agreement was achieved using the solvent viscosity, in agreement with the theoretical predictions for sterically interacting globular colloidal particles. Finally, the SANS results obtained at 24 degrees C were compared to the micelle self-diffusion coefficients previously measured by means of fluorescence correlation spectroscopy (FCS) at this temperature. The good agreement obtained after scaling the data with solution viscosity supports the validity of the generalized Stokes-Einstein relation in sterically interacting systems: the product of the colloidal particle self-diffusion coefficient and the macroscopic viscosity remains constant in a broad range of concentrations. It has been concluded that the FCS technique in combination with simple viscosity measurements might serve as a tool for estimating the micellar size and shape.

Evaporation into vacuum: Mass flux from momentum flux and the Hertz–Knudsen relation revisited

We performed molecular dynamics simulations of liquid film evaporation into vacuum for two cases: free evaporation without external supply of energy and evaporation at constant average liquid temperature. In both cases we found that the pressure inside a liquid film was constant, while temperature decreased and density increased as a function of distance from the middle of the film. The momentum flux in the vapor far from the liquid was equal to the liquid pressure in the evaporating film. Moreover the pseudopressure (stagnation pressure) was found to be constant in the evaporating vapor and equal to the liquid pressure. The momentum flux and its relation to the pressure determined the number of evaporating molecules per unit time and as a consequence the mass evaporation flux. We found a simple formula for the evaporation flux, which much better describes simulation results than the commonly used Hertz-Knudsen relation.

Close-packed monolayers of charged Janus-type nanoparticles at the air–water interface

We present a new method to obtain close-packed monolayers composed of noble metal nanoparticles (NP) possessing well-defined permanent charge of either sign. The method is based on the fact that ligands forming the protecting layer exhibit ability to rearrange at the NPs surface. We demonstrate that if the protecting layer is composed of a mixture of hydrophobic and hydrophilic charged ligands in appropriate proportion, the NP exhibits properties of a Janus-type particle with one of the hemispheres hydrophilic and the other hydrophobic. Such amphiphilic NPs self-assemble into a monolayer of well defined surface charge at the air-water interface. Due to strong stabilizing effect of the lateral electrostatic repulsions, such monolayer can be compressed to form close-packed hexagonal structure, and then easily transferred onto a solid substrate with the Langmuir-Blodgett technique.