Facilitated Diffusion Process Research Articles

Theoretical analysis of facilitated diffusion process in a liquid membrane: Adomian decomposition method

The mathematical model of facilitated transport systems in steady-state conditions is discussed. The nonlinear component associated with chemical reactions and diffusion in nonlinear equations forms the basis of this model. By solving the nonlinear equation using the Adomian decomposition method, we can obtain the analytical expression of the concentration of species. The simple expression of the facilitation factor ( ratio between the total flux of solute and that of the absence of carrier) is also reported. The parameters' influence on species concentration and facilitation factors are discussed. A good agreement is obtained when we compare the numerical results for all parameter values with new analytical results.

Optimizing bimetallic and multimetallic Cu-based catalysts by promoters for enhanced reverse Water-Gas Shift reaction: Insights and stability assessments

This study investigates bimetallic and multimetallic catalysts tailored for the reverse water gas shift (RWGS) reaction. Promoting with potassium and iron, while maintaining a constant copper content of 15 wt% on γ-alumina, revealed that bimetallic catalysts containing 7.5 wt% iron and 5 wt% potassium exhibit superior catalytic activity and stability. Among the multimetallic catalysts, the optimal composition was found to be in the 15 wt% Cu-5 wt% Fe-2.5 wt% K/γ-Al2O3 catalyst for the RWGS reaction. The prepared catalysts underwent assessments using various techniques such as N2 adsorption-desorption, XRD, H2-TPR, CO2-TPD, and FESEM. Stability assessments conducted over 72 hours showcased sustained long-term performance for the optimized catalysts, highlighting the influential role of the hierarchical structure. Mesopores extended the catalyst lifetime by reducing permeation limits, while macropores facilitated diffusion process, enhancing selectivity and stability. This hierarchical design also improved mass transfer, amplifying reaction rates. The synthesized catalysts exhibited exceptional specificity, demonstrating 100 % selectivity for the RWGS reaction. Particularly, under H2/CO2=1, these catalysts displayed remarkable carbon dioxide conversion rates, indicating potential for enhancing Cu-based catalysts in RWGS reactions by maximizing active sites.

Probing the modulation in facilitated diffusion guided by DNA-protein interactions in target search processes.

Many fundamental biophysical processes involving gene regulation and gene editing rely, at the molecular level, on an intricate methodology of searching and locating the precise target base pair sequence on the genome by specific binding proteins. A unique mechanism, known as 'facilitated diffusion', which is a combination of 1D sliding along with 3D movement, is considered to be the key step for such events. This also explains the relatively much shorter timescale of the target searching process, compared to other diffusion-controlled biophysical processes. In this work, we aim to probe the modulation of target search dynamics of a protein moiety by estimating the rate of the target search process, and the statistics of the search rounds and timescales accomplished by the 1D and 3D motions, based on first passage time (FPT) calculations. This is studied with its characteristics getting influenced by various given conditions such as, when the DNA is rigid or flexible, and when the target is placed at different locations on the DNA. The current theoretical framework includes a Brownian dynamics simulation setup adopting a straightforward coarse-grained model for a diffusing protein on DNA. Moreover, this theoretical analysis provides insights into the complex target search dynamics by highlighting the significance of the chain dynamics in the mechanistic details of the facilitated diffusion process.

Transcriptional factors control their diffusion on DNA by modulating their dynamics

Gene regulation requires transcription factors to efficiently recognize their target DNA site. Efficient recognition of target sequence is achieved by a facilitated diffusion process that combines 3D diffusion and 1D diffusive motions along the DNA, which are enabled by weak electrostatic (non-specific) interactions. The protein is presumed to scan the DNA while bound non-specifically, and lock into the specific binding site once it’s encountered. The mechanism by which a single domain protein alternates rapidly between these two binding modes is however unknown.

Facilitated Diffusion of Proline across Membranes of Liposomes and Living Cells by a Calix[4]pyrrole Cavitand

Summary The transport of anions and cations through liposomal membranes facilitated by synthetic carriers has been widely described. In contrast, analogous studies describing the facilitated transport of amino acids (aa) are scarce. We describe the use of calix[4]pyrrole receptors as synthetic carriers for the facilitated diffusion of aa across membranes of liposomes and living cells. We demonstrate that a calix[4]pyrrole cavitand is highly effective and selective for the facilitated diffusion of L-proline (L-Pro). We propose a mobile carrier diffusion mechanism to explain the observed aa facilitated diffusion process. The transport process involves the formation of a 1:1 complex between the carrier and the aa (e.g., L-Pro⊂1). We also describe the unprecedented application of a synthetic carrier to contribute to the uptake of L-Pro in human cells in addition to that mediated by the natural transporters. The reported results augur well for the potential use of aa synthetic carriers in therapeutic applications.

Facilitated Diffusion of Proline Across Membranes of Liposomes and Living Cells by a Calix[4]Pyrrole Cavitand

The transport of anions and cations through liposomal membranes facilitated by synthetic carriers has been widely described. In contrast, analogous studies describing the facilitated transport of amino acids (aa) are scarce. We describe the use of calix[4]pyrrole receptors as synthetic carriers for the facilitated diffusion of aa across membranes of liposomes and living cells. We demonstrate that a calix[4]pyrrole cavitand is highly effective and selective for the facilitated diffusion of L-proline (L-Pro). We propose a mobile carrier diffusion mechanism to explain the observed aa facilitated diffusion process. The transport process involves the formation of a 1:1 complex between the carrier and the aa (e.g. L-Pro⊂1). We also describe the unprecedented application of a synthetic carrier to contribute to the uptake of L-Pro in human cells in addition to that mediated by the natural transporters. The reported results augur well for the potential use of aa synthetic carriers in therapeutic applications.

Target search of a protein on DNA in the presence of position-dependent bias

We study the target search on DNA for proteins in the presence of non-constant drift. This search is realized by the facilitated diffusion process. Existing works on this problem focus on the case of constant drift. Starting from a non-local Fokker–Planck equation with the -order fractional Laplace operator and a ‘sink’ term, we obtain the possibility density function for a protein occurring at position x at time t. Based on this, we further compute the survival probability and the first arrival density in order to quantify the searching mechanisms. The numerical results show that in the linear drift case, there is an optimal index for the search to be most likely successful (searching reliability reaches its maximum). This optimal index depends on the initial position–target separation. It is also found that the diffusion intensity plays a positive role in improving the search success. The nonlinear double-well drift could drive the protein to reach the target with a larger possibility than the linear drag at the initial time period, but viewed over a long time duration, the linear drift is more beneficial for target search success. In contrast to the linear drift case, the search reliability and efficiency with nonlinear double-well drift have a monotonic relationship with the index, that is, the smaller the index is, the higher the likelihood of a protein finding its target.

Facilitated Diffusion Mechanisms in DNA Base Excision Repair and Transcriptional Activation.

Preservation of the coding potential of the genome and highly regulated gene expression over the life span of a human are two fundamental requirements of life. These processes require the action of repair enzymes or transcription factors that efficiently recognize specific sites of DNA damage or transcriptional regulation within a restricted time frame of the cell cycle or metabolism. A failure of these systems to act results in accumulated mutations, metabolic dysfunction, and disease. Despite the multifactorial complexity of cellular DNA repair and transcriptional regulation, both processes share a fundamental physical requirement that the proteins must rapidly diffuse to their specific DNA-binding sites that are embedded within the context of a vastly greater number of nonspecific DNA-binding sites. Superimposed on the needle-in-the-haystack problem is the complex nature of the cellular environment, which contains such high concentrations of macromolecules that the time frame for diffusion is expected to be severely extended as compared to dilute solution. Here we critically review the mechanisms for how these proteins solve the needle-in-the-haystack problem and how the effects of cellular macromolecular crowding can enhance facilitated diffusion processes. We restrict the review to human proteins that use stochastic, thermally driven site-recognition mechanisms, and we specifically exclude systems involving energy cofactors or circular DNA clamps. Our scope includes ensemble and single-molecule studies of the past decade or so, with an emphasis on connecting experimental observations to biological function.

Acceleration of bursty multiprotein target search kinetics on DNA by colocalisation.

Proteins are capable of locating specific targets on DNA by employing a facilitated diffusion process with intermittent 1D and 3D search steps. Gene colocalisation and coregulation-i.e. the spatial proximity of two communicating genes-is one factor capable of accelerating the target search process along the DNA. We perform Monte Carlo computer simulations and demonstrate the benefits of gene colocalisation for minimising the search time in a model DNA-protein system. We use a simple diffusion model to mimic the search for targets by proteins, produced initially in bursts of multiple proteins and performing the first-passage search on the DNA chain. The behaviour of the mean first-passage times to the target is studied as a function of distance between the initial position of proteins and the DNA target position, as well as versus the concentration of proteins. We also examine the properties of bursty target search kinetics for varying physical-chemical protein-DNA binding affinity. Our findings underline the relevance of colocalisation of production and binding sites for protein search inside biological cells.

Identification and Functional Characterization of a Novel OprD-like Chitin Uptake Channel in Non-chitinolytic Bacteria

Chitoporin from the chitinolytic marine Vibrio has been characterized as a trimeric OmpC-like channel responsible for effective chitin uptake. In this study we describe the identification and characterization of a novel OprD-like chitoporin (so-called EcChiP) from Escherichia coli The gene was identified, cloned, and functionally expressed in the Omp-deficient E. coli BL21 (Omp8) Rosetta strain. On size exclusion chromatography, EcChiP had an apparent native molecular mass of 50 kDa, as predicted by amino acid sequencing and mass analysis, confirming that the protein is a monomer. Black lipid membrane reconstitution demonstrated that EcChiP could readily form stable, monomeric channels in artificial phospholipid membranes, with an average single channel conductance of 0.55 ± 0.01 nanosiemens and a slight preference for cations. Single EcChiP channels showed strong specificity, interacting with long chain chitooligosaccharides but not with maltooligosaccharides. Liposome swelling assays indicated the bulk permeation of neutral monosaccharides and showed the size exclusion limit of EcChiP to be ∼200-300 Da for small permeants that pass through by general diffusion while allowing long chain chitooligosaccharides to pass through by a facilitated diffusion process. Taking E. coli as a model, we offer the first evidence that non-chitinolytic bacteria can activate a quiescent ChiP gene to express a functional chitoporin, enabling them to take up chitooligosaccharides for metabolism as an immediate source of energy.

Facilitated diffusion in the presence of obstacles on the DNA.

Biological functions of DNA depend on the sequence-specific binding of DNA-binding proteins to their corresponding binding sites. Binding of these proteins to their binding sites occurs through a facilitated diffusion process that combines three-dimensional diffusion in the cytoplasm with one-dimensional diffusion (sliding) along the DNA. In this work, we use a lattice model of facilitated diffusion to study how the dynamics of binding of a protein to a specific site (e.g., binding of an RNA polymerase to a promoter or of a transcription factor to its operator site) is affected by the presence of other proteins bound to the DNA, which act as 'obstacles' in the sliding process. Different types of these obstacles with different dynamics are implemented. While all types impair facilitated diffusion, the extent of the hindrance depends on the type of obstacle. As a consequence of hindrance by obstacles, more excursions into the cytoplasm are required for optimal target binding compared to the case without obstacles.

Permeability of the Blood-Brain Barrier: Molecular Mechanism of Transport of Drugs and Physiologically Important Compounds.

A new molecular model for the permeability of drugs and other physiologically important compounds to cross the blood-brain barrier has been developed. Permeability (logPS) is dependant on desolvation, lipophilicity, molecular volume and dipole moment. Previous models for BBB permeability have not considered desolvation and dipole moment as critical factors. The model applies to passive diffusion processes, and some facilitated diffusion processes. Passive permeability models may not apply to active transport processes, where complex membrane protein binding processes (e.g. stereoselectivity) are involved. Model phosphatidylcholine lipid bilayer membranes have been used to evaluate how charged or polar neutral compounds can interact through their molecular dipoles with the cell membrane to induce electromechanical changes in the cell membrane which facilitate permeation. The free energy of solvation in n-octanol has been shown to be a good measure of membrane lipophilicity by calculating the solvation free energy of a model PC lipid membrane in a series of closely related alcohols. The passive diffusion model for alcohols correlates with the known modulation of membrane bilayers which showed a size-dependent "cut-off" point in potency. For most drugs and related molecules, the neutral species are the permeating species.

The “Monkey‐Bar” Mechanism for Searching for the DNA Target Site: The Molecular Determinants

AbstractDNA recognition by DNA‐binding proteins, which is a pivotal event in most gene regulatory processes, is often preceded by an extensive search for the correct site. A facilitated diffusion process, in which a DBP combines 3D diffusion in solution with 1D sliding along DNA, has been suggested to explain how proteins can locate their target sites on DNA much faster than predicted by 3D diffusion alone. One of the key mechanisms in the localization of the target by a DNA‐binding protein is intersegment transfer in which the protein forms a bridged intermediate between two distant DNA regions. This jumping mechanism is more enhanced when the DNA‐binding protein is asymmetric in its structure or its dynamics. We suggest that asymmetry supports the “monkey bar” mechanism, in which different domains of the protein interact with different DNA regions. In this minireview, we discuss how the molecular architectures of the proteins and DNA may modulate the efficiency of monkey bar dynamics.

Carrier-mediated uptake of nobiletin, a citrus polymethoxyflavonoid, in human intestinal Caco-2 cells

The mechanism of intestinal absorption of nobiletin (NBL) was investigated using Caco-2 cells. The uptake of NBL from the apical membranes of Caco-2 cells was rapid and temperature-dependent and the presence of metabolic inhibitors, NaN3 and carbonylcyanide p-trifluoromethoxyphenylhydrazone, did not cause a decrease in NBL uptake. The relationship between the initial uptake of NBL and its concentration was saturable, suggesting the involvement of a carrier-mediated process. The Km and uptake clearance (Vmax/Km) values for NBL were 50.6 and 168.1μl/mg protein/min, respectively. This clearance value was about 9-fold greater than that of the non-saturable uptake clearance (Kd: 18.5μl/mg protein/min). The presence of structurally similar compounds, such as quercetin and luteolin, competitively inhibited NBL uptake. These results suggest that uptake of NBL from the apical membranes of Caco-2 cells is mainly mediated by an energy-independent facilitated diffusion process.

Dimethyl Ether Carbonylation to Methyl Acetate over Nanosized Mordenites

Nanosized mordenites were found to show significantly enhanced reaction efficiency in dimethyl ether (DME) carbonylation to methyl acetate (MA) because of a greatly facilitated diffusion process. Copper incorporation into the channels of the nanosized mordenites further promoted the reaction rate, selectivity, and stability. Moreover, upon the addition of a small amount of H2 (5–19 vol %) to the feed gas, deactivation was suppressed during DME carbonylation, whereas the catalyst stability and rate of formation of MA increased.

Obstacles May Facilitate and Direct DNA Search by Proteins

DNA recognition by DNA-binding proteins (DBPs), which is a pivotal event in most gene regulatory processes, is often preceded by an extensive search for the correct site. A facilitated diffusion process in which a DBP combines three-dimensional diffusion in solution with one-dimensional sliding along DNA has been suggested to explain how proteins can locate their target sites on DNA much faster than predicted by three-dimensional diffusion alone. Although experimental and theoretical studies have recently advanced understanding of the biophysical principles underlying the search mechanism, the process under in vivo cellular conditions is poorly understood. In this study, we used various computational approaches to explore how the presence of obstacle proteins on the DNA influences search efficiency. At a low obstacle occupancy (i.e., when few obstacles occupy sites on the DNA), sliding by the searching DBP may be confined, which may impair search efficiency. The obstacles, however, can be bypassed during hopping events, and the number of bypasses is larger for higher obstacle occupancies. Dynamism on the part of the obstacles may even further facilitate search kinetics. Our study shows that the nature and efficiency of the search process may be governed not only by the intrinsic properties of the DBP and the salt concentration of the medium, but also by the in vivo association of DNA with other macromolecular obstacles, their location, and occupancy.

DNA Looping Induced by Tubular Confinement

DNA looping is essential for the function and maintenance of genetic information. At short lengths the cyclization rate is limited by the semiflexible properties of DNA, while at long distances it is limited by the rapidly declining probability of colocation of the anchoring sequences of the loop. We propose that a facilitated diffusion process increases the rate of cyclization for large loops. The maze-like nuclear environment then becomes a major impediment for the required colocation.We have investigated the kinetic evolution of DNA loops (48500 bp) induced by T4 ligase inside a nanofabricated channel system with a channel cross-section of 100x100 nm2, and a few micron channel length. We found that addition of the ligase profoundly alters the behavior of DNA. In particular, ligase acts to stabilize hairpin geometries in which the extended forward and backward arms of the hairpin scan past each other. From the linear density of DNA inside the channel, we deduce that the effective excluded volume vanishes upon addition of T4 ligase and ATP.We conclude that the two strands are effectively stapled together through a large number of weak bonds involving T4 ligase. The absence of a similarly strong transformation of DNA in free solution points towards the necessity of parallel pre-alignment through the nanotube. We point to the fact that the formation of hairpins and the 1-d scanning of strands past each other solves the kinetic problem of forming a loop in a maze-like environment.

Dissection of the Xeroderma Pigmentosum Group C Protein Function by Site-Directed Mutagenesis

Xeroderma pigmentosum group C (XPC) protein is a sensor of helix-distorting DNA lesions, the function of which is to trigger the global genome repair (GGR) pathway. Previous studies demonstrated that XPC protein operates by detecting the single-stranded character of non-hydrogen-bonded bases opposing lesion sites. This mode of action is supported by structural analyses of the yeast Rad4 homologue that identified critical side chains making close contacts with a pair of extrahelical nucleotides. Here, alanine substitutions of the respective conserved residues (N754, F756, F797, F799) in human XPC were tested for DNA-binding activity, accumulation in tracks and foci of DNA lesions, nuclear protein mobility, and the induction of downstream GGR reactions. This study discloses a dynamic interplay between XPC protein and DNA, whereby the association with one displaced nucleotide in the undamaged strand mediates the initial encounter with lesion sites. The additional flipping-out of an adjacent nucleotide is necessary to hand over the damaged site to the next GGR player. Surprisingly, this mutagenesis analysis also reveals that the rapid intranuclear trafficking of XPC protein depends on constitutive interactions with native DNA, implying that the search for base damage takes place in living cells by a facilitated diffusion process.

Glucose transport from mother to fetus—A theoretical study

The factors that affect and govern the glucose transfer from maternal blood to the fetus are not completely deciphered. We present a steady state, one dimensional mathematical simulation which integrates the main mechanisms that have been shown to exist: metabolic consumption of the placenta, simple and facilitated diffusion via the two membranes of the microvillous and simple diffusion within the placenta. The model uses all available physiologic data we could collect. Numerical results indicate that the most crucial factor in determining the fetal glucose concentration is the facilitated diffusion process at the basal membrane or, more specifically: the permeability of the basal membrane and the density of the transporter GLUT1 on its faces. The gradient between the maternal and the fetal glucose concentration is important as is the metabolic consumption of the placenta. The diffusion within the placenta and the conditions that prevail at the apical microvillous plasma membrane are much less significant. Intrasyncytial concentration of glucose is close to that of maternal blood. The adjustment of the fetal glucose concentration to abrupt changes of its surrounding is estimated to be quite rapid hence for all practical purposes this steady state model can serve as a reasonable approximation. Parameters that await experimental determination are identified.

Recycling of Dolichyl Monophosphate to the Cytoplasmic Leaflet of the Endoplasmic Reticulum after the Cleavage of Dolichyl Pyrophosphate on the Lumenal Monolayer

During protein N-glycosylation, dolichyl pyrophosphate (Dol-P-P) is discharged in the lumenal monolayer of the endoplasmic reticulum (ER). Dol-P-P is then cleaved to Dol-P by Dol-P-P phosphatase (DPPase). Studies with the yeast mutant cwh8Delta, lacking DPPase activity, indicate that recycling of Dol-P produced by DPPase contributes significantly to the pool of Dol-P utilized for lipid intermediate biosynthesis on the cytoplasmic leaflet. Whether Dol-P formed in the lumen diffuses directly back to the cytoplasmic leaflet or is first dephosphorylated to dolichol has not been determined. Incubation of sealed ER vesicles from calf brain with acetyl-Asn-Tyr-Thr-NH(2), an N-glycosylatable peptide, to generate Dol-P-P in the lumenal monolayer produced corresponding increases in the rates of Man-P-Dol, Glc-P-Dol, and GlcNAc-P-P-Dol synthesis in the absence of CTP. No changes in dolichol kinase activity were observed. When streptolysin-O permeabilized CHO cells were incubated with an acceptor peptide, N-glycopeptide synthesis, requiring multiple cycles of the dolichol pathway, occurred in the absence of CTP. The results obtained with sealed microsomes and CHO cells indicate that Dol-P, formed from Dol-P-P, returns to the cytoplasmic leaflet where it can be reutilized for lipid intermediate biosynthesis, and dolichol kinase is not required for recycling. It is possible that the flip-flopping of the carrier lipid is mediated by a flippase, which would provide a mechanism for the recycling of Dol-P derived from Man-P-Dol-mediated reactions in N-, O-, and C-mannosylation of proteins, GPI anchor assembly, and the three Glc-P-Dol-mediated reactions in Glc(3)Man(9)GlcNAc(2)-P-P-Dol (DLO) biosynthesis.