Complex Morphology of the Intermediate Phase in Block Copolymers and Semicrystalline Polymers As Revealed by ¹H NMR Spin Diffusion Experiments

Nanostructured multiphase polymers exhibiting a mobile and a rigid phase also contain a phase of intermediate mobility that is usually assumed to be a continuous, uninterrupted interphase layer. This assumption is contrary to recent molecular-resolution micrographs and contradicts results from NMR spin diffusion experiments, all of which suggest a nontrivial interface structure. In this contribution, we reconsider our previous ¹H NMR spin diffusion data sets (Roos Colloid. Polym. Sci. 2014, 292, 1825) and perform optimized 2D and 3D numerical spin diffusion calculations to characterize the basic intermediate-phase morphological pattern, thus overcoming previous inconsistencies in data fitting. For the diblock copolymer poly­(butadiene)-poly­(styrene), PS<i>-<i>b</i>-</i>PB, we demonstrate that the interphase region comprises nanometer-size intermixed immobile, intermediate and mobile subregions. In contrast, for the semicrystalline polymer poly­(ε-caprolactone), PCL, the spin diffusion data are best reproduced by an intermediate phase that is fully embedded within the rigid phase, which is attributed to an imperfect crystalline structure. For both samples, the new findings reveal a complex discontinuous, dynamically inhomogeneous structure of the intermediate phase

Fast Magic-Angle-Spinning ¹⁹F Spin Exchange NMR for Determining Nanometer ¹⁹F–¹⁹F Distances in Proteins and Pharmaceutical Compounds

Internuclear distances measured using NMR provide crucial constraints of three-dimensional structures but are often restricted to about 5 Å due to the weakness of nuclear-spin dipolar couplings. For studying macromolecular assemblies in biology and materials science, distance constraints beyond 1 nm will be extremely valuable. Here we present an extensive and quantitative analysis of the feasibility of ¹⁹F spin exchange NMR for precise and robust measurements of interatomic distances up to 1.6 nm at a magnetic field of 14.1 T, under 20–40 kHz magic-angle spinning (MAS). The measured distances are comparable to those achievable from paramagnetic relaxation enhancement but have higher precision, which is better than ±1 Å for short distances and ±2 Å for long distances. For ¹⁹F spins with the same isotropic chemical shift but different anisotropic chemical shifts, intermediate MAS frequencies of 15–25 kHz without ¹H irradiation accelerate spin exchange. For spectrally resolved ¹⁹F–¹⁹F spin exchange, ¹H–¹⁹F dipolar recoupling significantly speeds up ¹⁹F–¹⁹F spin exchange. On the basis of data from five fluorinated synthetic, pharmaceutical, and biological compounds, we obtained two general curves for spin exchange between CF groups and between CF₃ and CF groups. These curves allow ¹⁹F–¹⁹F distances to be extracted from the measured spin exchange rates after taking into account ¹⁹F chemical shifts. These results demonstrate the robustness of ¹⁹F spin exchange NMR for distance measurements in a wide range of biological and chemical systems

Coupling and Decoupling of Rotational and Translational Diffusion of Proteins under Crowding Conditions

Molecular motion of biopolymers <i>in vivo</i> is known to be strongly influenced by the high concentration of organic matter inside cells, usually referred to as crowding conditions. To elucidate the effect of intermolecular interactions on Brownian motion of proteins, we performed ¹H pulsed-field gradient NMR and fluorescence correlation spectroscopy (FCS) experiments combined with small-angle X-ray scattering (SAXS) and viscosity measurements for three proteins, αB-crystalline (αBc), bovine serum albumin, and hen egg-white lysozyme (HEWL) in aqueous solution. Our results demonstrate that long-time translational diffusion quantitatively follows the expected increase of macro-viscosity upon increasing the protein concentration in all cases, while rotational diffusion as assessed by polarized FCS and previous multi-frequency ¹H NMR relaxometry experiments reveals protein-specific behavior spanning the full range between the limiting cases of full decoupling from (αBc) and full coupling to (HEWL) the macro-viscosity. SAXS was used to study the interactions between the proteins in solution, whereby it is shown that the three cases cover the range between a weakly interacting hard-sphere system (αBc) and screened Coulomb repulsion combined with short-range attraction (HEWL). Our results, as well as insights from the recent literature, suggest that the unusual rotational–translational coupling may be due to anisotropic interactions originating from hydrodynamic shape effects combined with high charge and possibly a patchy charge distribution