A nanomechanical interface to rapid single-molecule interactions

Abstract

single-molecule techniques provide opportunities for molecularly precise imaging, manipulation, assembly and biophysical studies. owing to the kinetics of bond rupture processes, rapid single-molecule measurements can reveal novel bond rupture mechanisms, probe singlemolecule events with short lifetimes and enhance the interaction forces supplied by single molecules. Rapid measurements will also increase throughput necessary for technological use of single-molecule techniques. Here we report a nanomechanical sensor that allows single-molecule force spectroscopy on the previously unexplored microsecond timescale. We probed bond lifetimes around 5 µs and observed significant enhancements in molecular interaction forces. our loading-rate-dependent measurements provide experimental evidence for an additional energy barrier in the biotin-streptavidin complex. We also demonstrate quantitative mapping of rapid single-molecule interactions with high spatial resolution. This nanomechanical interface may allow studies of molecular processes with short lifetimes and development of novel biological imaging, single-molecule manipulation and assembly technologies.

Coexistence of ribbon and helical fibrils originating from hIAPP 20–29 revealed by quantitative nanomechanical atomic force microscopy

“…High-resolution solidstate NMR structures of hIAPP 20-29 fibrils have provided detailed insight into the antiparallel hetero zipper with a twist along the fibril axis (21-23). However, no uniform detailed theory of the self-assembly behavior is available (11) and, in particular, the transitions between different intermediates during fibrillation remain to be elucidated.Here, we apply high-resolution AFM and the recently developed microsecond force spectroscopy (μFS) for quantitative nanomechanical maps (24, 25) to explore the nanostructures and nanomechanical properties of species formed during the self-assembly of the decapeptide hIAPP [20][21][22][23][24][25][26][27][28][29] . By following the temporal evolution of the amyloid peptide self-assembly process, we calculate the average thickening speed of ribbon structures, which is considered a key intermediate in the ribbon-like packing scheme (26,27).…”

“…Transitions among the various intermediate stages are the subject of many studies but are not yet fully elucidated. Here, we combine high-resolution atomic force microscopy and quantitative nanomechanical mapping to determine the self-assembled structures of the decapeptide hIAPP [20][21][22][23][24][25][26][27][28][29] , which is considered to be the fibrillating core fragment of the human islet amyloid polypeptide (hIAPP) involved in type-2 diabetes. We successfully follow the evolution of hIAPP [20][21][22][23][24][25][26][27][28][29] nanostructures over time, calculate the average thickening speed of small ribbon-like structures, and provide evidence of the coexistence of ribbon and helical fibrils, highlighting a key step within the self-assembly model.…”

“…Here, we combine high-resolution atomic force microscopy and quantitative nanomechanical mapping to determine the self-assembled structures of the decapeptide hIAPP [20][21][22][23][24][25][26][27][28][29] , which is considered to be the fibrillating core fragment of the human islet amyloid polypeptide (hIAPP) involved in type-2 diabetes. We successfully follow the evolution of hIAPP [20][21][22][23][24][25][26][27][28][29] nanostructures over time, calculate the average thickening speed of small ribbon-like structures, and provide evidence of the coexistence of ribbon and helical fibrils, highlighting a key step within the self-assembly model. In addition, the mutations of individual side chains of wide-type hIAPP [20][21][22][23][24][25][26][27][28][29] shift this balance and destabilize the helical fibrils sufficiently relative to the twisted ribbons to lead to their complete elimination.…”

“…We successfully follow the evolution of hIAPP [20][21][22][23][24][25][26][27][28][29] nanostructures over time, calculate the average thickening speed of small ribbon-like structures, and provide evidence of the coexistence of ribbon and helical fibrils, highlighting a key step within the self-assembly model. In addition, the mutations of individual side chains of wide-type hIAPP [20][21][22][23][24][25][26][27][28][29] shift this balance and destabilize the helical fibrils sufficiently relative to the twisted ribbons to lead to their complete elimination. We combine atomic force microscopy structures, mechanical properties, and solid-state NMR structural information to build a molecular model containing β sheets in cross-β motifs as the basis of selfassembled amyloids.…”

“…hIAPP is a 37-amino-acid peptide hormone, and the decapeptide SNNFGAILSS comprising hIAPP [20][21][22][23][24][25][26][27][28][29] , considered to be the core fibrillating element of hIAPP relating to T2D (18,19), is critical for the fibrillation of hIAPP and shows cytotoxicity (19). The analysis of hIAPP [20][21][22][23][24][25][26][27][28][29] fibrils revealed two distinct fibril types, comprising either parallel β-strands or antiparallel β-strands (20). High-resolution solidstate NMR structures of hIAPP [20][21][22][23][24][25][26][27][28][29] fibrils have provided detailed insight into the antiparallel hetero zipper with a twist along the fibril axis (21)(22)(23).…”