Niitsu, A. | Search bioRxiv

bioRxiv2024

De novo design of α-helical peptide channels with designer stoichiometry

Despite advances in peptide and protein design, the rational design of membrane-spanning peptides that form conducting channels remains challenging due to our imperfect understanding of the sequence-to-structure relationships that drive membrane insertion, assembly, and conductance. Here, we describe the design and computational and experimental characterization of a series of coiled coil-based peptides that form transmembrane -helical barrels. Through a combination of rational and computational design, we obtain barrels with 5 to 7 helices, as characterized in detergent micelles. In lipid bilayers, these peptide assemblies exhibit two conductance states with relative populations dependent on the applied potential: (i) a low-conductance states that correlate with variations in the modeled coiled-coil barrel geometries, indicating stable transmembrane -helical barrels; and (ii) high-conductance states in which single pores change size in discrete steps. Notably, the high-conductance states are similar for all peptides in contrast to the low-conductance states. This indicates the formation of large, dynamic pores through the recruitment and expulsion of peptides, as observed in natural barrel-stave peptide pores. These findings establish rational routes to design and tune functional membrane-spanning peptide channels with specific conductance and geometry.

synthetic biology↗

bioRxiv2022

Repulsive interaction and secondary structure of highly charged proteins in regulating biomolecular condensation

Biomolecular condensation is involved in various cellular processes both functional and dysfunctional. Regulation of the condensation is thus crucial to avoid pathological protein aggregation and to maintain stable cellular environments. Recently, a class of highly charged intrinsically disordered proteins (IDPs), which are called the heat-resistant obscure (Hero) proteins, are shown to protect other client proteins from pathological aggregation. Besides the potential importance of this function, molecular mechanisms for how Hero proteins can protect other proteins from aggregation are not still known. Here we perform multiscale molecular dynamics (MD) simulations of Hero11, one of the Hero proteins, and the C-terminal region of TDP-43, as a target protein of Hero11, at various conditions to examine how they interact with each other. Based on the simulation results, three possible mechanisms have been proposed: (i) TDP-43 and Hero11 in dense phase reduces contacts with each other and shows faster diffusion due to the repulsive Hero11-Hero11 interactions, (ii) the amount of TDP-43 in dilute phase increases and their sizes become greater upon the attractive Hero11-TDP-43 interactions, and (iii) Hero-11 on the surface of small TDP-43 condensates avoids their fusions with the repulsive interactions. We also examine possible Hero-11 structures in atomistic and coarse-grained MD simulations and found disordered Hero-11 tend to assemble on the surface of the condensates, avoiding the droplet fusion effectively. The proposed mechanisms give us new insight into the regulation of biomolecular condensation in the cells and other conditions.

biophysics↗