Hfsp Journal最新文献

Transmissible spongiform encephalopathies (TSEs) are lethal infectious neurodegenerative diseases. TSEs are caused by prions, infectious agents lacking informational nucleic acids, and possibly identical with higher-order aggregates of the cellular glycolipoprotein PrP(C). Prion strains are derived from TSE isolates that, even after inoculation into genetically identical hosts, cause disease with distinct patterns of protein aggregate deposition, incubation times, morphology of the characteristic brain damage, and cellular tropism. Most of these traits are relatively stable across serial passages. Here we review current techniques for studying prion strain differences in vivo and in cells, and discuss the strain phenomena in the general context of the knowledge gained from modeling prion fibril growth in vitro and in simple organisms.

The universe of conformational substates of a protein molecule is huge. The complete energy landscape of proteins is, therefore, complex when studied at low temperature. Many experiments under physiological conditions commonly reveal a simpler spectrum of states. These states are individually ensembles of low temperature substates. That is, room temperature experiments probe the low free energy part of the spectrum of excitations. This paper describes how the complete landscape and the spectrum of these thermally excited motions can be related to each other. On funneled landscapes, partially folded ensembles of states are the most important excited states. Their properties and their free energy spectrum can often be predicted by native topology based models. Frustration, i.e., the conflict between inconsistent stabilizing interactions that have evolved for other purposes than optimizing folding, offers another mechanism for forming low free energy excitations. Frustration can be localized and quantified using energy landscape theory. Symmetry provides an obvious route to low free energy states in oligomeric systems, where simply repositioning parts of the molecule in ways quasi-equivalent to their relation in the native structure gives nearly degenerate energies.

Great advances have been made in electron microscopy (EM) over the past decade, with the result that a number of protein complexes have been solved at near-atomic resolution using EM imaging. However, only a limited number of such complexes are expected to have the high degree of internal order needed to achieve this type of resolution. Many other complexes and polymers will be visualized and reconstructed by EM at an intermediate level of resolution, where the polypeptide chain cannot be directly traced. Crystal and nuclear magnetic resonance structures for components or subunits of these higher-order assemblies are frequently available. One of the greatest strengths of EM continues to be the ability to dock high-resolution structures of components into low or intermediate resolution reconstructions of assemblies to build pseudoatomic models for quaternary structure. This review discusses the strengths and limitations of this approach, with particular emphasis on protein polymers. I discuss how limitations in resolution can lead to ambiguities in building models, and these cannot be always be resolved with available data. The use of homology models for quaternary structure are particularly problematic, given accumulating evidence for the divergence of quaternary structures at the same time that tertiary structure can be conserved.

The role of water in biomolecule dynamics has attracted much interest over the past decade, due in part to new probes of biomolecule-water interactions and developments in molecular simulations. Terahertz (THz) spectroscopy, among the most recent experimental methods brought to bear on this problem, is able to detect even small solute induced changes of the collective water network dynamics at the biomolecule-water interface. THz measurements reveal that proteins influence up to 1000 water molecules in their surroundings, and that even small saccharides influence the dynamics of hundreds of surrounding water molecules. The THz spectrum of a protein is sensitive to mutation and depends on the surface charge and flexibility of the protein. Influence on the solvation shell appears most pronounced for native wildtype proteins and decreases upon partial unfolding or mutation. THz spectra of solvated saccharides reveal that the number of water molecules coupled dynamically to a saccharide, forming a dynamical hydration shell around it, is related to the number of exposed oxygen atoms on the solute. The thickness of this layer appears correlated with the bioprotection efficiency of the saccharide. All findings support the thesis of a long-range dynamic coupling between biomolecule and solvent.

The dynamic nature of protein function is a fundamental concept in the physics of proteins. Although the basic general ideas are well accepted most experimental evidence has an indirect nature. The detailed characterization of the dynamics is necessary for the understanding in detail. The dynamic fluctuations thought crucial for the function span an extremely broad time, starting from the picosecond regime. Recently, a few new experimental techniques emerged that permit the observation of dynamical phenomena directly. Notably, pulsed infrared (IR) spectroscopy has been applied with great success to observe structural changes with picosecond time resolution. Using two-dimensional-IR vibrational echo chemical exchange spectroscopy Ishikawa and co-workers [Ishikawa et al. (2008), Proc. Natl. Acad. Sci. U.S.A. 101, 14402-14407] managed to observe the transition between well defined conformational substrates of carbonmonoxy myoglobin directly. This is an important step in improving our insight into the details of protein function.

Knowledge of a protein's three-dimensional native structure is vital in determining its chemical properties and functionality. However, experimental methods to determine structure are very costly and time-consuming. Computational approaches such as folding simulations and structure prediction algorithms are quicker and cheaper but lack consistent accuracy. This currently restricts extensive computational studies to abstract protein models. It is thus essential that simplifications induced by the models do not negate scientific value. Key to this is the use of thoroughly defined proteinlike sequences. In such cases abstract models can allow for the investigation of important biological questions. Here, we present a procedure to generate and classify proteinlike sequence data sets. Our LatPack tools and the approach in general are applicable to arbitrary lattice protein models. Identification is based on thermodynamic kinetic features and incorporates the sequential assembly of proteins by addressing cotranslational folding. We demonstrate the approach in the widely used unrestricted 3D-cubic HP-model. The resulting sequence set is the first large data set for this model exhibiting the proteinlike properties required. Our data tools are freely available and can be used to investigate protein-related problems.

The unfolded state ensemble of proteins has been described as a structurally featureless state. While this approach is supported by the fact that many unfolded proteins follow the scaling law behavior of a random coil, there is evidence that the unfolded states of various proteins are stabilized by native or non-native interactions. Recently, the existence of extensive non-native structure was reported for a repeat protein, which resulted in a scaling law exponent that is significantly smaller than that of a random polymer [Cortajarena et al., J. Mol. Biol. 382(1), 203-212 (2008)]. It was concluded that the high compactness of this protein stems from a significant fraction of interacting PP(II) helical segments in the unfolded state. In this study, we aim at providing possible molecular understanding of this anomalous compactness of the unfolded state and to investigate its origin. Using a hierarchy of computational models, we ask whether in general the unfolded state of a repeat protein is likely to be intrinsically more compact than the unfolded state of globular proteins, or whether this phenomenon depends mostly on the occurrence of a specific sequence that promotes PP(II) conformations. Our results suggest that the formation of the PP(II) conformation is indeed essential, yet the recurring sequence of repeat proteins promotes the interactions between these PP(II) segments and the formation of non-native interactions in the unfolded state.

Green fluorescent protein (GFP) is a large protein with a complex eleven-stranded beta-barrel structure. Previous studies have shown that it has a complex energy landscape for folding on which there are several intermediate states and a denatured state with significant residual structure. Here, we use two different types of HD exchange measurement and nuclear magnetic resonance (NMR) techniques to probe the energy landscape for folding of GFP in further detail. HD exchange experiments were performed over a wide range of conditions including different concentrations of denaturant. Results show that the penetration model dominates the exchange mechanism, consistent with the known stability and slow unfolding kinetics of GFP. HD exchange experiments at high pH establish that there is an extremely slow-exchanging superstable core of amide protons in GFP that are clustered and located in beta-strands 1, 2, 4, 5, and 6. These residues form part of a mini-beta-sheet which we propose constitutes a folding nucleus. Using a pulsed-labeling strategy, the acid-denatured state has been investigated and the residual structure observed in earlier studies shown to locate to beta-strands 1 and 3. There is some evidence that this residual structure is stabilized by a localized hydrophobic collapse of the polypeptide chain.

Genetic recombination in eukaryotes requires the pairing of homologous chromosomes to allow precise molecular exchanges between chromosome pairs at intertwined structures called Holliday junctions, the formation of which requires the action of the RecA protein. The mechanism behind the precise pairing of structures as long as chromosomes remains mysterious. In yeast, during the initial phases of meiosis, chromosomes are paired at approximately 65 kilobase intervals via paranemic interactions that do not involve strand breakage nor the intervention of analogs of the RecA protein. It has been proposed that these paranemic interactions could occur between G-rich chromosomal regions, but putting in register stretches of homologous sequences hundreds of kb long remains challenging. Recent developments on the theory of the physicochemical properties of DNA in aqueous solutions, in presence of di- or multivalent counterions, leads to the prediction that molecules with the same sequence tend to pair spontaneously by paranemic interactions depending on the electrostatic properties of DNA. Experimental support for this prediction has now been provided in vitro with naked DNA. This newly discovered property of DNA duplexes may thus provide a clue to solve the puzzle of the premeiotic pairing.