Molecular and Ionic Diffusion in Ion Exchange Membranes and Biological Systems (Cells and Proteins) Studied by NMR.

The results of NMR, and especially pulsed field gradient NMR (PFG NMR) investigations, are summarized. Pulsed field gradient NMR technique makes it possible to investigate directly the partial self-diffusion processes in spatial scales from tenth micron to millimeters. Modern NMR spectrometer diffusive units enable to measure self-diffusion coefficients from 10-13 m2/s to 10-8 m2/s in different materials on 1 H, 2 H, 7 Li, 13 C, 19 F, 23 Na, 31 P, 133 Cs nuclei. PFG NMR became the method of choice for reveals of transport mechanism in polymeric electrolytes for lithium batteries and fuel cells. Second wide field of application this technique is the exchange processes and lateral diffusion in biological cells as well as molecular association of proteins. In this case a permeability, cell size, and associate lifetime could be estimated. The authors have presented the review of their research carried out in Karpov Institute of Physical Chemistry, Moscow, Russia; Institute of Problems of Chemical Physics RAS, Chernogolovka, Russia; Kazan Federal University, Kazan, Russia; Korea University, Seoul, South Korea; Yokohama National University, Yokohama, Japan. The results of water molecule and Li+, Na+, Cs+ cation self-diffusion in Nafion membranes and membranes based on sulfonated polystyrene, water (and water soluble) fullerene derivative permeability in RBC, casein molecule association have being discussed.

On Complex Formation between 5-Fluorouracil and ß-Cyclodextrin in Solution and in the Solid State: IR Markers and Detection of Short-Lived Complexes by Diffusion NMR.

In this work, the nuclear magnetic resonance (NMR) and IR spectroscopic markers of the complexation between 5-fluorouracil (5-FU) and ß-cyclodextrin (ß-CD) in solid state and in aqueous solution are investigated. In the attenuated total reflectance(ATR) spectra of 5-FU/ß-CD products obtained by physical mixing, kneading and co-precipitation, we have identified the two most promising marker bands that could be used to detect complex formations: the C=O and C-F stretching bands of 5-FU that experience a blue shift by ca. 8 and 2 cm-1 upon complexation. The aqueous solutions were studied by NMR spectroscopy. As routine NMR spectra did not show any signs of complexation, we have analyzed the diffusion attenuation of spin-echo signals and the dependence of the population factor of slowly diffusing components on the diffusion time (diffusion NMR of pulsed-field gradient (PFG) NMR). The analysis has revealed that, at each moment, ~60% of 5-FU molecules form a complex with ß-CD and its lifetime is ca. 13.5 ms. It is likely to be an inclusion complex, judging from the independence of the diffusion coefficient of ß-CD on complexation. The obtained results could be important for future attempts of finding better methods of targeted anticancer drug delivery.

Translational diffusion of unfolded and intrinsically disordered proteins.

Translational (or self-diffusion) coefficient in dilute solution is inversely proportional to the size of a diffusing molecule, and hence self-diffusion coefficient measurements have been applied to determine the effective hydrodynamic radii for a range of native and nonnative protein conformations. In particular, translational diffusion coefficient measurements are useful to estimate the hydrodynamic radius of natively (or intrinsically) disordered proteins in solution, and, thereby, probe the compactness of a protein as well as its change when environmental parameters such as temperature, solution pH, or protein concentration are varied. The situation becomes more complicated in concentrated solutions. In this review, we discuss the translational diffusion of disordered proteins in dilute and crowded solutions, focusing primarily on the information provided by pulsed-field gradient NMR technique, and draw analogies to well-structured globular proteins and synthetic polymers.

Effect of Reducing Agent TCEP on Translational Diffusion and Supramolecular Assembly in Aqueous Solutions of α-Casein.

The translational diffusion coefficient is highly sensitive to the size change of diffusing species and is ideally suited for the study of molecular association. Here, we used translational diffusion measurements by a pulsed-field gradient nuclear magnetic resonance (PFG NMR) technique to investigate the role of disulfide bonds in the formation of a supramolecular gel-like structure in the concentrated solution of α-casein. To reduce disulfide bonds, we added a commonly used reducing reagent tris(2-carboxyethyl)phosphine (TCEP) to α-casein solution. We found that the disruption of a disulfide bond Cys36-Cys40 in αs2-casein does not alter the translational diffusion or secondary structure of α-casein in dilute, 1 and 3% (wt %) solution. On the contrary, in concentrated, 15% (wt %) α-casein solution, in addition to the disruption of disulfide bonds, TCEP induced significant changes in gel properties. New long-lived intermolecular interactions formed, leading to the irreversible gel formation. While a few side reactions of TCEP (as well as other reducing agents, e.g., dithiothreitol) have been reported, this area is still understudied. Here, we provide new data on the side reaction of the reducing agent TCEP in concentrated protein solution, suggesting that at high protein concentrations TCEP should be used with caution.

Effect of Intrinsic Disorder and Self-Association on the Translational Diffusion of Proteins: The Case of α-Casein.

Translational diffusion is the major mode of macromolecular transport in leaving organisms, and therefore it is vital to many biological and biotechnological processes. Although translational diffusion of proteins has received considerable theoretical and experimental scrutiny, much of that attention has been directed toward the description of globular proteins. The translational diffusion of intrinsically disordered proteins (IDPs), however, is much less studied. Here, we use a pulsed-gradient nuclear magnetic resonance technique (PFG NMR) to investigate the translational diffusion of a disordered protein in a wide range of concentrations using α-casein that belongs to the class of natively disordered proteins as an example.