Conformational Ensemble of Monomeric α-Synuclein in Aqueous and Crowded Environments as revealed by Markov State Model

Abstract

140-residue intrinsically disordered protein α-synuclein (αS) is known to be susceptible to environmental cues/crowders and adopts conformations that are vastly variable in the extent of secondary structure and tertiary interactions. Depending upon the nature of these interactions, some of the conformations may be suitable for its physiological functions while some may be predisposed to aggregate with other partners into higher ordered species or to phase separate. However, the inherently heterogenous and dynamic nature of αS has precluded a clear demarcation of its monomeric precursor between aggregation-prone and functionally relevant aggregation-resistant states. Here, we optimally characterize a set of metastable conformations of αS by developing a comprehensive Markov state model (MSM) using cumulative 108 μs-long all-atom MD simulation trajectories of monomeric αS. Notably, the dimension of the most populated metastable (85%) state (Rg ∼ 2.59 (±0.45) nm) corroborates PRE-NMR studies of αS monomer and undergoes kinetic transition at 0.1-150 μs time-scale with weakly populated (0.06%) random-coil like ensemble (Rg ∼ 5.85 (±0.43) nm) and globular protein-like state (14%) (Rg ∼ 1.95 (±0.08) nm). The inter-residue contact maps identify a set of mutually interconverting aggregation-prone β-sheet networks in the NAC region and aggregation-resistant long-range interactions between N- and C-terminus or helical conformations. The presence of crowding agents compacts the MSM-derived metastable conformations in a non-monotonic fashion and skews the ensemble by either introducing new tertiary contacts or reinforcing the innate contacts to adjust to the excluded-volume effects of such environments. These observations of crucial monomeric states would serve as important steps towards rationalising routes that trigger αS-associated pathologies.