posted on 2021-08-10, 16:08authored byJitao Wen, Liu Hong, Georg Krainer, Qiong-Qiong Yao, Tuomas P. J. Knowles, Si Wu, Sarah Perrett
Liquid–liquid
phase separation (LLPS) of proteins into biomolecular
condensates has emerged as a fundamental principle underpinning cellular
function and malfunction. Indeed, many human pathologies, including
protein misfolding diseases, are linked to aberrant liquid-to-solid
phase transitions, and disease-associated protein aggregates often
nucleate through phase separation. The molecular level determinants
that promote pathological phase transitions remain, however, poorly
understood. Here we study LLPS of the microtubule-associated protein
Tau, whose aberrant aggregation is associated with a number of neurodegenerative
diseases, including Alzheimer’s disease. Using single molecule
spectroscopy, we probe directly the conformational changes that the
protein undergoes as a result of LLPS. We perform single-molecule
FRET and fluorescence correlation spectroscopy experiments to monitor
the intra- and intermolecular changes and demonstrate that the N-
and C-terminal regions of Tau become extended, thus exposing the microtubule-binding
region. These changes facilitate intermolecular interactions and allow
for the formation of nanoscale clusters of Tau. Our results suggest
that these clusters can promote the fibrillization of Tau, which can
be dramatically accelerated by disease-related mutations P301L and
P301S. Our findings thus provide important molecular insights into
the mechanism of protein phase separation and the conversion of protein
condensates from functional liquid assemblies to pathological aggregates.