170 Nanometer Nuclear Magnetic Resonance Imaging using Magnetic Resonance Force Microscopy

We demonstrate one-dimensional nuclear magnetic resonance imaging of the semiconductor GaAs with 170 nanometer slice separation and resolve two regions of reduced nuclear spin polarization density separated by only 500 nanometers. This is achieved by force detection of the magnetic resonance, Magnetic Resonance Force Microscopy (MRFM), in combination with optical pumping to increase the nuclear spin polarization. Optical pumping of the GaAs creates spin polarization up to 12 times larger than the thermal nuclear spin polarization at 5 K and 4 T. The experiment is sensitive to sample volumes containing $\sim 4 \times 10^{11}$ $^{71}$Ga$/\sqrt{Hz}$. These results demonstrate the ability of force-detected magnetic resonance to apply magnetic resonance imaging to semiconductor devices and other nanostructures.Comment: Submitted to J of Magnetic Resonanc

Successive stages of amyloid-? self-assembly characterized by solid-state nuclear magnetic resonance with dynamic nuclear polarization

Self-assembly of amyloid-β (Aβ) peptides in human brain tissue leads to neurodegeneration in Alzheimer’s disease (AD). Amyloid fibrils, whose structures have been extensively characterized by solid state nuclear magnetic resonance (ssNMR) and other methods, are the thermodynamic end point of Aβ self-assembly. Oligomeric and protofibrillar assemblies, whose structures are less well-understood, are also observed as intermediates in the assembly process in vitro and have been implicated as important neurotoxic species in AD. We report experiments in which the structural evolution of 40-residue Aβ (Aβ40) is monitored by ssNMR measurements on frozen solutions prepared at four successive stages of the self-assembly process. Measurements on transient intermediates are enabled by ssNMR signal enhancements from dynamic nuclear polarization (DNP) at temperatures below 30 K. DNP-enhanced ssNMR data reveal a monotonic increase in conformational order from an initial state comprised primarily of monomers and small oligomers in solution at high pH, to larger oligomers near neutral pH, to metastable protofibrils, and finally to fibrils. Surprisingly, the predominant molecular conformation, indicated by ¹³C NMR chemical shifts and by side chain contacts between F19 and L34 residues, is qualitatively similar at all stages. However, the in-register parallel β-sheet supramolecular structure, indicated by intermolecular ¹³C spin polarization transfers, does not develop before the fibril stage. This work represents the first application of DNP-enhanced ssNMR to the characterization of peptide or protein self-assembly intermediates

Structure of FUS Protein Fibrils and Its Relevance to Self-Assembly and Phase Separation of Low-Complexity Domains

Polymerization and phase separation of proteins containing low-complexity (LC) domains are important factors in gene expression, mRNA processing and trafficking, and localization of translation. We have used solid-state nuclear magnetic resonance methods to characterize the molecular structure of self-assembling fibrils formed by the LC domain of the fused in sarcoma (FUS) RNA-binding protein. From the 214-residue LC domain of FUS (FUS-LC), a segment of only 57 residues forms the fibril core, while other segments remain dynamically disordered. Unlike pathogenic amyloid fibrils, FUS-LC fibrils lack hydrophobic interactions within the core and are not polymorphic at the molecular structural level. Phosphorylation of core-forming residues by DNA-dependent protein kinase blocks binding of soluble FUS-LC to FUS-LC hydrogels and dissolves phase-separated, liquid-like FUS-LC droplets. These studies offer a structural basis for understanding LC domain self-assembly, phase separation, and regulation by post-translational modification