Biology subjects

Ritsch, I.

Publications and source records attributed to Ritsch, I..

3 recordsLinked to original sources

bioRxiv2026

From Hummingbird to Elephant: Amyloid Formation in Natural Transthyretin Variants

Transthyretin (TTR) is a secreted protein associated with cardiac and other amyloid diseases via misfolding. We have previously shown that agitation of human TTR solutions at neutral pH results in aggregation and fibril formation. Here we report that agitation-induced aggregation of TTR from species with very different heart rates (Annas hummingbird, hbTTR, and African elephant, aeTTR) differs from that of human TTR (huTTR). Aggregation of hbTTR is slow and favors formation of smaller, fibrillar aggregates, while aeTTR aggregation is rapid and favors larger, more amorphous particles. Spherical, early-stage oligomeric intermediates were found for all variants by mass photometry and electron microscopy. The slow aggregation of hbTTR matches its resistance to denaturation by 8 M urea. The widely different aggregation behavior exhibited by these naturally occurring TTR variants in response to mechanical agitation under close to physiological conditions provides insight into how small sequence differences can contribute to the evolutionary fitness of different animals.

biophysics↗

bioRxiv2024

Aggregation of Transthyretin by Fluid Agitation

The transthyretin (TTR) tetramer, assembled as a dimer of dimers, transports thyroxine and retinol binding protein in blood plasma and cerebrospinal fluid. Aggregation of wild type or pathogenic variant TTR leads to transthyretin amyloidosis (ATTR), which is associated with neurodegenerative and cardiac disease. The trigger for TTR aggregation under physiological conditions is unknown. The tetramer is extremely stable at neutral pH, but aggregation via tetramer dissociation and monomer misfolding can be induced in vitro by lowering the pH. To elucidate factors that may cause TTR aggregation at neutral pH, we examined the effect of shear forces such as arise from fluid flow in the vascular system. Fluid shear forces were generated by rapidly stirring TTR solutions in conical microcentrifuge tubes. Under agitation, TTR formed {beta}-rich aggregates and fibrils at a rate that was dependent upon protein concentration. The lag time before the onset of agitation-induced aggregation increases as the total TTR concentration is increased, consistent with a mechanism in which the tetramer first dissociates to form monomer that either partially unfolds to enter the aggregation pathway or reassociates to form tetramer. NMR spectra recorded at various time points during the lag phase revealed growth of an aggregation-prone intermediate trapped as a dynamically perturbed tetramer. Enhanced conformational fluctuations in the weak dimer-dimer interface suggests loosening of critical inter-subunit contacts which likely destabilizes the agitated tetramer and predisposes it towards dissociation. These studies provide new insights into the mechanism of aggregation of wild type human TTR under near physiological conditions.

biophysics↗

bioRxiv2022

Phase separation of hnRNP A1 upon specific RNA-binding observed by magnetic resonance

Interaction of heterogeneous nuclear ribonucleoprotein A1 (hnRNP A1) with specific single-stranded RNA and its relation to liquid-liquid phase separation were investigated in vitro by magnetic resonance based on site-directed spin labelling. An ensemble model of free hnRNP A1 in the absence of RNA was derived from distance distributions between spin labelled sites and small angle X-ray scattering. This model revealed a compact state of the low-complexity domain and interaction of this domain with the RNA recognition motifs. Paramagnetic relaxation enhancement NMR spectroscopy confirmed this interaction. The addition of RNA to dispersed solutions of hnRNP A1 induced phase separation, observed by formation of liquid droplets. The phase separation depended on the RNA concentration and sequence, with continuous wave EPR spectroscopy showing that local protein dynamics is affected by point mutations in the RNA sequence. We propose that an interplay of sequence-specific RNA binding and phase transition serves as a regulatory mechanism for RNA segregation in the stress response of cells.

molecular biology↗