Molecular Insights into the Dynamics of Amyloid Fibril Growth: Elongation and Lateral Assembly of GNNQQNY Protofibrils.

The self-assembly of peptides and proteins into ß-sheet rich amyloid fibrils is linked to both functional and pathological states. In this study, the growth of fibrillar structures of the short peptide GNNQQNY, a fragment from the yeast prion Sup35 protein, was examined. Molecular dynamics simulations were used to study alternative mechanisms of fibril growth, including elongation through binding of monomers as well as fibril self-assembly into larger, more mature structures. It was found that after binding, monomers diffused along preformed fibrils toward the ends, supporting the mechanism of fibril growth via elongation. Lateral assembly of protofibrils was found to occur readily, suggesting that this could be the key to transitioning from isolated fibrils to mature multilayer structures. Overall, the work provides mechanistic insights into the competitive pathways that govern amyloid fibril growth.

Simulations of the c-subunit of ATP-synthase reveal helix rearrangements.

The c-subunit of the enzyme, ATP synthase couples the proton movement through the a-subunit with its own rotation and subsequent rotation of the F1 ring to drive ATP synthesis. Here, we perform mus time-scale coarse-grained molecular dynamics simulations of the c-subunit to characterize its structure and dynamics. Two different helix-helix interfaces, albeit with similar interfacial characteristics, are sampled in the simulations. The helix-2 of the c-subunit monomer rotates around the axis of helix-1 bringing about a change in the interface. Previous models have also proposed such a change in the helix interface but postulated that helix-2 swivels around its own axis. Such large-scale changes in helix packing motifs have not been observed before. The helix-swirling persists even in the c-subunit ring but the dynamics is much slower. The cooperative behavior in the ring appears to stabilize a conformation less-populated in the monomer. Analyzing the stability of the c-subunit ring, it was found that six lipid molecules are necessary to fill the central cavity of the ring. These lipid molecules were not aligned with the surrounding bilayer but protruded towards the periplasmic side. The characterization of the monomer and ring presented in this work sheds light into the structural dynamics of the c-subunit and its functional relevance.

Factors that affect the degree of twist in beta-sheet structures: a molecular dynamics simulation study of a cross-beta filament of the GNNQQNY peptide.

By exploiting the recent availability of the crystal structure of a cross-beta filament of the GNNQQNY peptide fragment of the yeast prion protein Sup35, possible factors affecting the twisting of beta-sheets structures have been analyzed. The advantage of this system is that it is composed entirely of beta-sheet and thus free of potential ambiguities present in systems studied previously. In the crystal the cross-beta filament consists of antiparallel beta-sheets formed by parallel and in register peptides lying perpendicular to the long axis of the filament. The results of a series of molecular dynamics simulations performed under different conditions indicate that in the absence of crystal packing interactions there is no free energy barrier against twisting for the cross-beta filament found planar in the crystal. More specifically, we find that there is only a small change in enthalpy (<3 kJ mol(-1) per residue) for twists in the range 0-12 degrees with the planar form (in the crystal environment) being enthalpically stabilized. In contrast, entropic contributions, in particular those associated with an increase in backbone dynamics upon twisting, stabilize the twisted form. The degree of twist was found to vary depending on the environmental conditions as the result from an apparent subtle interplay of multiple small contributions. These observations are consistent with the different degrees of twist observed in beta-sheets both in native protein structures and amyloid fibrils.

Pathogenic mutations shift the equilibria of alpha-synuclein single molecules towards structured conformers.

Alpha-synuclein (alpha-Syn) is an abundant brain protein whose mutations have been linked to early-onset Parkinson's disease (PD). We recently demonstrated, by means of a single-molecule force spectroscopy (SMFS) methodology, that the conformational equilibrium of monomeric wild-type (WT) alpha-Syn shifts toward beta-containing structures in several unrelated conditions linked to PD pathogenicity. Herein, we follow the same methodology previously employed for WT alpha-Syn to characterize the conformational heterogeneity of pathological alpha-Syn mutants A30P, A53T, and E46K. Contrary to the bulk ensemble-averaged spectroscopies so far employed to this end by different authors, our single-molecule methodology monitored marked differences in the conformational behaviors of the mutants with respect to the WT sequence. We found that all the mutants have a much higher propensity than the WT to adopt a monomeric compact conformation that is compatible with the acquiring of beta structure. Mutants A30P and A53T show a similar conformational equilibrium that is significantly different from that of E46K. Another class of conformations, stabilized by mechanically weak interactions (MWI), shows a higher variety in the mutants than in the WT protein. In the A30P mutant these interactions are relatively stronger, and therefore the corresponding conformations are possibly more structured. The more structured and globular conformations of the mutants can explain their higher propensity to aggregate with respect to the WT.

Structure of spheroidal HDL particles revealed by combined atomistic and coarse-grained simulations.

Spheroidal high-density lipoprotein (HDL) particles circulating in the blood are formed through an enzymatic process activated by apoA-I, leading to the esterification of cholesterol, which creates a hydrophobic core of cholesteryl ester molecules in the middle of the discoidal phospholipid bilayer. In this study, we investigated the conformation of apoA-I in model spheroidal HDL (ms-HDL) particles using both atomistic and coarse-grained molecular dynamics simulations, which are found to provide consistent results for all HDL properties we studied. The observed small contribution of cholesteryl oleate molecules to the solvent-accessible surface area of the entire ms-HDL particle indicates that palmitoyloleoylphosphatidylcholines and apoA-I molecules cover the hydrophobic core comprised of cholesteryl esters particularly well. The ms-HDL particles are found to form a prolate ellipsoidal shape, with sizes consistent with experimental results. Large rigid domains and low mobility of the protein are seen in all the simulations. Additionally, the average number of contacts of cholesteryl ester molecules with apoA-I residues indicates that cholesteryl esters interact with protein residues mainly through their cholesterol moiety. We propose that the interaction of annular cholesteryl oleate molecules contributes to apoA-I rigidity stabilizing and regulating the structure and function of the ms-HDL particle.

Topological properties of the energy landscape of small peptides.

In several works it has been shown that the interplay between short range and long range interactions, mimicking the hydrophobic effect, leads to the formation of the typical secondary structures in proteins, alpha-helices and beta-sheets. In this work we study in detail how the general properties of the energy landscape emerge in a model that presents both components. In this regard it proves useful a sort of perturbative approach. In our model many features of the energy landscape in absence of long range interaction can be determined analytically. The comparison between the energy landscape of this reduced model to that of the complete model gives interesting insight on the role of long range interactions.