Diffusion of protein

Johnson et al. [186] measured diffusion of fluorescein-labeled macromolecules in agarose gels. Their data agreed well with Eq. (85), which combined the hydrodynamic effects with the steric hindrance factors. Gibbs and Johnson [131] measured diffusion of proteins and smaller molecules in polyacrylamide gels using pulsed-field gradient NMR methods and found their data to fit the stretched exponential form... 

Rate of protein transfer to or from a reverse micellar phase and factors affecting the rate are important for the practical applications of RME for the extraction and purification of proteins/enzymes and for scale-up. The mechanism of protein exchange between two immiscible phases (Fig. 2) can be divided into three steps [36] the diffusion of protein from bulk aqueous solution to the interface, the formation of a protein-containing micelle at the interface, and the diffusion of a protein-containing micelle in to the organic phase. The reverse steps are applicable for back transfer with the coalescence of protein-filled RM with the interface to release the protein. The overall mass transfer rate during an extraction processes will depend on which of these steps is rate limiting. 

As indicated in my report, we now know the rates of lateral diffusion of phospholipids in lipid bilayers in the fluid state, and in a few cases the rates of lateral diffusion of proteins in fluid lipids are also known. At the present time nothing is known about the rates of lateral diffusion of phospholipids in the crystalline, solid phases of the substances. As mentioned in my report, there are reasons to suspect that the rates of lateral diffusion of phospholipids in the solid solution crystalline phases of binary mixtures of phospholipids may be appreciable on the experimental time scale. Professor Ubbelohde may well be correct in pointing out the possibility of diffusion caused by defects. However, such defects, if present, apparently do not lead to significant loss of the membrane permeability barrier, except at domain boundaries. 

The expected greater size of protein-polysaccharide complexes can reduce the diffusion rate of the adsorbing species towards the interface. This effect is especially important for small monomeric proteins. In addition, Ganzevles and co-workers (2006) have suggested that the diffusion of protein in the complexes may not solely be responsible for the slow surface tension decay. Rather, the gradual dissociation (and subsequent adsorption) of protein from complexes, when they are in close proximity to the interface, could also contribute to the behaviour. 

The central event in the development of liver fibrosis is the enhanced sinusoidal deposition of extracellular matrix proteins that are mainly produced by activated HSC [86, 112, 113] and to a minor extent by endothelial cells [44-46] and hepatocytes [114, 115]. So far, no evidence has been found that KC are directly involved in the production of extracellular matrix proteins [39]. The accumulation of extracellular matrix proteins is caused by a disturbed balance between the synthesis and the degradation of the matrix proteins. This imbalance leads to a 5 to 10-fold increase in the total amount of matrix molecules and to an altered composition of the extracellular matrix. In contrast to normal livers, the sinusoids in fibrotic livers are stuffed with the fibrillar collagens type I and III. This colla-genization of the sinusoids, referred to as sinusoidal capillarization, causes severe disturbances of the blood flow and an impaired exchange of proteins between the liver cells and blood. Furthermore, this capillarization is accompanied by a loss of fenestration of the sinusoidal endothelial lining, which further hampers the diffusion of proteins between plasma and hepatic cells. 

A. H. Stolpen, D. E. Golan, and J. S. Pober, Tumor necrosis factor and immune interferon act in concert to slow the lateral diffusion of proteins and lipids in human endothelial cell membranes, J. Cell Biol. 707 781-789 (1988). 

Separation of proteins on the basis of size can be achieved by means of gel filtration. This technique relies on diffusion of protein molecules into the pores of a gel matrix in a column. A commonly used type of gel material is dextran, a polymer of glucose, in the form of very small beads. This material is available commercially as Sephadex in a range of different pore sizes. 

Eventually, this method allows quantitative measuring of the translational diffusion of proteins modified with these three labels in solution and in biomembranes. The minimal approximate volume of a sample available for the fluorescence measurement (using a regular commercial spectrofluorimeter) in this method is about 10"3 pi when the total concentration of fluorophores is close to 0.01 pM and the local concentration of radicals is about 10 pM. 

Since CSF is in direct contact with the environment of the central nervous system (C NS), it is obvious that any changes in biochemical composition of brain parenchyma should be predominantly reflected in the CSF. A recent review by Reiber (2001) presents a complete concept of distinguishing diffusion of brain-derived proteins into CSF from diffusion of proteins from blood into CSF, allowing proteins that originate in the brain to be prioritized. Lumbar pimcture is an easy procedure, with a low incidence of complications. In a large study (Andreasen et al, 2001), only 4.1% of all patients experienced postlumbar headache, and an even smaller proportion of 2% was reported by Blennow et al. (1993). Thus, it is reasonable to postulate that lumbar puncture (LP) is a feasible and only moderately invasive procedure, and that CSF analysis could possibly improve current clinical and neuroimaging-based approaches to diagnosis. 

Cell membranes are two-dimensional fluids that exhibit a wide range of dynamic behaviors. Recent technical advances have enabled unprecedented views of membrane dynamics in living cells. In this technical review, we provide a brief overview of three well-studied examples of membrane dynamics lateral diffusion of proteins and lipids in the plane of the membrane, vesicular trafficking between intracellular compartments, and exchange of proteins on and off membranes. We then discuss experimental approaches to monitor membrane protein and lipid dynamics, and we place a special emphasis on the use of genetically encoded fluorescent probes and live cell-imaging techniques. 

How rapidly diffusion occurs is characterized by the diffusion coefficient D, a parameter that provides a measure of the mean of the squared displacement x of a molecule per unit time f. For diffusion in two dimensions such as a membrane, this is given by = 4Ht. The Saffman-Delbrtlck model of Brownian motion in biologic membranes describes the relationship between membrane viscosity, solvent viscosity, the radius R and height of the diffusing species, and D for both lateral and rotational diffusion of proteins in membranes (3, 4). This model predicts for example that for lateral diffusion, D should be relatively insensitive to the radius of the diffusing species, scaling with log (1/R). 

Clegg RM, Vaz WLC. Translational diffusion of proteins and lipids in artificial lipid bilayer membranes. A comparison of experiment with theory. In Progress in Protein-Lipid Interactions, Vol. I. Watts A, De Pont JJHHM, eds. 1985. Elsevier, Amsterdam, The Netherlands, pp. 173-229... 

Lipids diffuse freely in fluid model membranes. FRAP measurements show full recovery and diffusion coefficients on the order of magnitude of 10 cm /sec. Free diffusion with a similar rate is often observed for lipids in the biomembrane. However, many cell membrane proteins show lower diffusion rates and incomplete recovery after photobleaching. For membrane proteins, dramatically different behavior in model and biological membranes is a common case. In model membranes, membrane proteins also diffuse freely and their diffusion coefficients are often similar to the diffusion coefficients of lipids. On the contrary, in biomembranes, the diffusion of proteins is 2-3 orders of magnitude slower and the fluorescence recovery is often incomplete. This observation points to limitations of the fluid mosaic model as will be discussed below. 

Another aspect of lipid-protein interaction that is conve-luently studied in supported bilayers is the lateral diffusion of proteins and lipids and their influence on each other. The regulatory lipid phosphatidyl-inositol-biphosphate (PIP2) slows the diffusion of syntaxin in supported bilayers (37). Conversely, increasing syntaxin concentrations decrease the diffusion of PIP2 and to a lesser extent that of phosphatidylserine. In another system, in which the transmembrane domain of the fibroblast growth factor receptor was incorporated into supported bilay-ers, lipid and protein diffusion were measured (60). Although protein diffusion was slow (0.006 ttm /s), lipid diffusion was fast (2.6 ttm /s). 

One-dimensional diffusion can accelerate the formation of site-specific interactions within biopolymers by up to lO -fold (Berg et al., 1981). Such facihtated diffusion is used by transcription factors and restriction endonucleases to locate specific sites on double-stranded DNA (von Hippel and Berg, 1989). The backbone of RNA, like that of DNA, could allow for the facilitated diffusion of proteins. Yet, the facilitated diffusion of a protein along RNA (or any single-stranded nucleic acid) has not been demonstrated previously. 

The diffusion of proteins and peptides in solution is dictated by the same considerations as those discussed in section 3.6. The rate of translational movement depends on the size of the molecule, its shape and interactions with solvent molecules. The rate of translational movement is often expressed by a frictional coefficient, f, defined in relation to the diffusion coefficient D, by equation (11.4) ... 

If staining is used to visualize the separated proteins, the proteins are usually first fixed by precipitating them in the gel with a chemical agent like acetic acid and methanol. This prevents diffusion of proteins out of the gel when submersed in the stain solution. The amount of dye taken up by the sample is affected by many factors, such as the type of protein and the degree of denaturation of the proteins by the fixing agents. 

The galactolipids are known to be distributed asymmetrically in the lipid bilayer - about 60% of the galactoUpids are present in the outer leaflets and 40% in the inner leaflets of the thylakoid-membrane bilayer. The strong interaction between the head-groups of the galactolipid molecules determine then-packing properties and enhance the stability of the membrane. Formation of the bilayer structure of the thylakoid membrane also depends on the presence of proteins. Another characteristic of the thylakoid lipids is their high content ofthe trienoic acid, (C18 3) a-linolenic acid, which contributes to the fluidity of the membrane, necessary for the diffusion of lipophilic compounds such as plastoquinone, and the lateral diffusion of protein complexes. 

To analyze the effect of emulsifier concentration and aqueous phase composition on the kinetics of diffusion (fcdifr)/ the evolution of the slope of tt vs. 9 or E vs. 9 is of great utility (Nino et al., 2003). We have observed that fcdiff increases with the emulsifier concentration in the bulk phase according to the Ward and Tordai equation. The presence of sucrose in the aqueous phase increases the diffusion of proteins towards the interface. However, is lower in ethanol aqueous solutions than in water. In addition, decreases with increasing ethanol or at higher sucrose concentrations in the aqueous phase (Nino et al., 2003). 

Problem 7-23. Diffusivity of a Protein in a Porous Medium. The diffusion of proteins in a porous medium, such as gels or chromatographic columns, is important for many processes in biotechnology. These include separations of valuable proteins, mass transfer in bioreactors, and biosensing. We wish to determine how the diffusivity of the protein scales with its size and the nature of the porous medium. 

Tight junctions block the diffusion of proteins and some lipids In the plane of the plasma membrane, contributing to the polarity of epithelial cells. They also limit and regulate the extracellular (paracellular) flow of water and solutes from one side of the epithelium to the other (see Figure 6-11). 

W6. Wunderly, C., Gel structure and diffusion of proteins. Clin. Chim. Acta 4, 754 (1959). 

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