Structural Model of the Proline-Rich Domain of Huntingtin Exon-1 Fibrils.

Huntington's disease is a heritable neurodegenerative disease that is caused by a CAG expansion in the first exon of the huntingtin gene. This expansion results in an elongated polyglutamine domain that increases the propensity of huntingtin exon-1 to form cross-ß fibrils. Although the polyglutamine domain is important for fibril formation, the dynamic, C-terminal proline-rich domain (PRD) of huntingtin exon-1 makes up a large fraction of the fibril surface. Because potential fibril toxicity has to be mediated by interactions of the fibril surface with its cellular environment, we wanted to model the conformational space adopted by the PRD. We ran 800-ns long molecular dynamics simulations of the PRD using an explicit water model optimized for intrinsically disordered proteins. These simulations accurately predicted our previous solid-state NMR data and newly acquired electron paramagnetic resonance double electron-electron resonance distances, lending confidence in their accuracy. The simulations show that the PRD generally forms an imperfect polyproline (polyP) II helical conformation. The two polyP regions within the PRD stay in a polyP II helix for most of the simulation, whereas occasional kinks in the proline-rich linker region cause an overall bend in the PRD structure. The dihedral angles of the glycine at the end of the second polyP region are very variable, effectively decoupling the highly dynamic 12 C-terminal residues from the rest of the PRD.

The complex role of the N-terminus and acidic residues of HdeA as pH-dependent switches in its chaperone function.

HdeA is a small acid-stress chaperone protein found in the periplasm of several pathogenic gram-negative bacteria. In neutral pH environments HdeA is an inactive folded homodimer but when exposed to strong acidic environments it partially unfolds and, once activated, binds to other periplasmic proteins, protecting them from irreversible aggregation. Here we use a combination of hydrogen/deuterium exchange NMR experiments and constant pH molecular dynamics simulations to elucidate the role of HdeA's N-terminus in its activation mechanism. Previous work indicates that the N-terminus is flexible and unprotected at high pH while exhibiting interactions with some HdeA client binding site residues. It, however, becomes partially solvent-protected at pH 2.6 - 2.8 and then loses protection again at pH 2.0. This protection is not due to the appearance of new secondary structure, but rather increased contacts between N-terminal residues and the C-terminus of the other protomer in the dimer, as well as concurrent loosening of its hold on the client binding site residues, priming HdeA for interactions with periplasmic client proteins. This work also uncovers unusual protonation profiles of some titratable residues and suggests their complex role in chaperone function.

PCAViz: An Open-Source Python/JavaScript Toolkit for Visualizing Molecular Dynamics Simulations in the Web Browser.

Molecular dynamics (MD) simulations reveal molecular motions at atomic resolution. Recent advances in high-performance computing now enable microsecond-long simulations capable of sampling a wide range of biologically relevant events. But the disk space required to store an MD trajectory increases with simulation length and system size, complicating collaborative sharing and visualization. To overcome these limitations, we created PCAViz, an open-source toolkit for sharing and visualizing MD trajectories via the web browser. PCAViz includes two components: the PCAViz Compressor, which compresses and saves simulation data; and the PCAViz Interpreter, which decompresses the data in users' browsers and feeds it to any of several browser-based molecular-visualization libraries (e.g., 3Dmol.js, NGL Viewer, etc.). An easy-to-install WordPress plugin enables "plug-and-play" trajectory visualization. PCAViz will appeal to a broad audience of researchers and educators. The source code is available at http://durrantlab.com/pcaviz/ , and the WordPress plugin is available via the official WordPress Plugin Directory.

HER3-targeted protein chimera forms endosomolytic capsomeres and self-assembles into stealth nucleocapsids for systemic tumor homing of RNA interference in vivo.

RNA interference represents a potent intervention for cancer treatment but requires a robust delivery agent for transporting gene-modulating molecules, such as small interfering RNAs (siRNAs). Although numerous molecular approaches for siRNA delivery are adequate in vitro, delivery to therapeutic targets in vivo is limited by payload integrity, cell targeting, efficient cell uptake, and membrane penetration. We constructed nonviral biomaterials to transport small nucleic acids to cell targets, including tumor cells, on the basis of the self-assembling and cell-penetrating activities of the adenovirus capsid penton base. Our recombinant penton base chimera contains polypeptide domains designed for noncovalent assembly with anionic molecules and tumor homing. Here, structural modeling, molecular dynamics simulations, and functional assays suggest that it forms pentameric units resembling viral capsomeres that assemble into larger capsid-like structures when combined with siRNA cargo. Pentamerization forms a barrel lined with charged residues mediating pH-responsive dissociation and exposing masked domains, providing insight on the endosomolytic mechanism. The therapeutic impact was examined on tumors expressing high levels of HER3/ErbB3 that are resistant to clinical inhibitors. Our findings suggest that our construct may utilize ligand mimicry to avoid host attack and target the siRNA to HER3+ tumors by forming multivalent capsid-like structures.