On-Pathway Oligomer of Human Islet Amyloid Polypeptide Induced and Stabilized by Mechanical Rotation during Magic Angle Spinning Nuclear Magnetic Resonance.

Intermediates along the fibrillation pathway are generally considered to be the toxic species responsible for the pathologies of amyloid diseases. However, structural studies of these species have been hampered by heterogeneity and poor stability under standard aqueous conditions. Here, we report a novel methodology for producing stable, on-pathway oligomers of the human type-2 diabetes-associated islet amyloid polypeptide (hIAPP or amylin) using the mechanical forces associated with magic angle spinning (MAS). The species were a heterogeneous mixture of globular and short rod-like species with significant ß-sheet content and the capability of seeding hIAPP fibrillation. We used MAS nuclear magnetic resonance to demonstrate that the nature of the species was sensitive to sample conditions, including peptide concentration, ionic strength, and buffer. The methodology should be suitable for studies of other aggregating systems.

On-Pathway Oligomer of Human Islet Amyloid Polypeptide Induced and Stabilized by Mechanical Rotation During MAS NMR.

Intermediates along the fibrillation pathway are generally considered to be the toxic species responsible for the pathologies of amyloid diseases. However, structural studies of these species have been hampered by heterogeneity and poor stability in standard aqueous conditions. Here, we report a novel methodology for producing stable, on-pathway oligomers of the human Type-2 Diabetes-associated islet amyloid polypeptide (hIAPP, or amylin) using the mechanical forces associated with magic angle spinning (MAS). The species were a heterogeneous mixture of globular and short rod-like species with significant beta-sheet content and the capability of seeding hIAPP fibrillation. We used MAS NMR to demonstrate that the nature of the species was sensitive to sample conditions including peptide concentration, ionic strength, and buffer. The methodology should be suitable for studies of other aggregating systems.

Atomic Resolution of Cotton Cellulose Structure Enabled by Dynamic Nuclear Polarization Solid-State NMR.

The insufficient resolution of conventional methods has long limited the structural elucidation of cellulose and its derivatives, especially for those with relatively low crystallinities or in native cell walls. Recent 2D/3D solid-state NMR studies of 13C uniformly labeled plant biomaterials have initiated a re-investigation of our existing knowledge in cellulose structure and its interactions with matrix polymers but for unlabeled materials, this spectroscopic method becomes impractical due to limitations in sensitivity. Here, we investigate the molecular structure of unlabeled cotton cellulose by combining natural abundance 13C-13C 2D correlation solid-state NMR spectroscopy, as enabled by the sensitivity-enhancing technique of dynamic nuclear polarization (DNP), with statistical analysis of the observed and literature-reported chemical shifts. The atomic resolution allows us to monitor the loss of Iα and Iß allomorphs and the generation of a novel structure during ball-milling, which reveals the importance of large crystallite size for maintaining the Iα and Iß model structures. Partial order has been identified in the "disordered" domains, as evidenced by a discrete distribution of well-resolved peaks. This study not only provides heretofore unavailable high-resolution insights into cotton cellulose but also presents a widely applicable strategy for analyzing the structure of cellulose-rich materials without isotope-labeling. This work was part of a multi-technique study of ball-milled cotton described in the previous article in the same issue.

Molecular architecture of fungal cell walls revealed by solid-state NMR.

The high mortality of invasive fungal infections, and the limited number and inefficacy of antifungals necessitate the development of new agents with novel mechanisms and targets. The fungal cell wall is a promising target as it contains polysaccharides absent in humans, however, its molecular structure remains elusive. Here we report the architecture of the cell walls in the pathogenic fungus Aspergillus fumigatus. Solid-state NMR spectroscopy, assisted by dynamic nuclear polarization and glycosyl linkage analysis, reveals that chitin and α-1,3-glucan build a hydrophobic scaffold that is surrounded by a hydrated matrix of diversely linked ß-glucans and capped by a dynamic layer of glycoproteins and α-1,3-glucan. The two-domain distribution of α-1,3-glucans signifies the dual functions of this molecule: contributing to cell wall rigidity and fungal virulence. This study provides a high-resolution model of fungal cell walls and serves as the basis for assessing drug response to promote the development of wall-targeted antifungals.