Helix-coil transition of PrP106-126: molecular dynamic study.

A set of 34 molecular dynamic (MD) simulations totaling 305 ns of simulation time of the prion protein-derived peptide PrP106-126 was performed with both explicit and implicit solvent models. The objective of these simulations is to investigate the relative stability of the alpha-helical conformation of the peptide and the mechanism for conversion from the helix to a random-coil structure. At neutral pH, the wild-type peptide was found to lose its initial helical structure very fast, within a few nanoseconds (ns) from the beginning of the simulations. The helix breaks up in the middle and then unwinds to the termini. The spontaneous transition into the random coil structure is governed by the hydrophobic interaction between His(111) and Val(122). The A117V mutation, which is linked to GSS disease, was found to destabilize the helix conformation of the peptide significantly, leading to a complete loss of helicity approximately 1 ns faster than in the wild-type. Furthermore, the A117V mutant exhibits a different mechanism for helix-coil conversion, wherein the helix begins to break up at the C-terminus and then gradually to unwind towards the N-terminus. In most simulations, the mutation was found to speed up the conversion through an additional hydrophobic interaction between Met(112) and the mutated residue Val(117), an interaction that did not exist in the wild-type peptide. Finally, the beta-sheet conformation of the wild-type peptide was found to be less stable at acidic pH due to a destabilization of the His(111)-Val(122), since at acidic pH this histidine is protonated and is unlikely to participate in hydrophobic interaction.

Modeling the 3D structure of GPCRs from sequence.

G-protein-coupled receptors (GPCRs) are a large and functionally diverse protein superfamily, which form a seven transmembrane (TM) helices bundle with alternating extra-cellular and intracellular loops. GPCRs are considered to be one of the most important groups of drug targets because they are involved in a broad range of body functions and processes and are related to major diseases. In this paper we present a new technology, named PREDICT, for modeling the 3D structure of any GPCR from its amino acid sequence. This approach takes into account both internal protein properties (i.e., the amino acid sequence) and the properties of the membrane environment. Unlike competing approaches, the new technology does not rely on the single known structure of rhodopsin, and is thus capable of predicting novel GPCR conformations. We demonstrate the capabilities of PREDICT in reproducing the known experimental structure of rhodopsin. In principle, PREDICT-generated models offer new opportunities for structure-based drug discovery towards GPCR targets.

Evidence for novel functions of the keratin tail emerging from a mutation causing ichthyosis hystrix.

Unraveling the molecular basis of inherited disorders of epithelial fragility has led to understanding of the complex structure and function of keratin intermediate filaments. Keratins are organized as a central alpha-helical rod domain flanked by nonhelical, variable end domains. Pathogenic mutations in 19 different keratin genes have been identified in sequences corresponding to conserved regions at the beginning and end of the rod. These areas have been recognized as zones of overlap between aligned keratin proteins and are thought to be crucial for proper assembly of keratin intermediate filaments. Consequently, all keratin disorders of skin, hair, nail, and mucous membranes caused by mutations in rod domain sequences are characterized by perinuclear clumping of fragmented keratin intermediate filaments, thus compromising mechanical strength and cell integrity. We report here the first mutation in a keratin gene (KRT1) that affects the variable tail domain (V2) and results in a profoundly different abnormality of the cytoskeletal architecture leading to a severe form of epidermal hyperkeratosis known as ichthyosis hystrix Curth-Macklin. Structural analyses disclosed a failure in keratin intermediate filament bundling, retraction of the cytoskeleton from the nucleus, and failed translocation of loricrin to the desmosomal plaques. These data provide the first in vivo evidence for the crucial role of a keratin tail domain in supramolecular keratin intermediate filament organization and barrier formation.

Solvent effects on the energy landscapes and folding kinetics of polyalanine.

The effect of a solvation on the thermodynamics and kinetics of polyalanine (Ala(12)) is explored on the basis of its energy landscapes in vacuum and in an aqueous solution. Both energy landscapes are characterized by two basins, one associated with alpha-helical structures and the other with coil and beta-structures of the peptide. In both environments, the basin that corresponds to the alpha-helical structure is considerably narrower than the basin corresponding to the beta-state, reflecting their different contributions to the entropy of the peptide. In vacuum, the alpha-helical state of Ala(12) constitutes the native state, in agreement with common helical propensity scales, whereas in the aqueous medium, the alpha-helical state is destabilized, and the beta-state becomes the native state. Thus solvation has a dramatic effect on the energy landscape of this peptide, resulting in an inverted stability of the two states. Different folding and unfolding time scales for Ala(12) in hydrophilic and hydrophobic chemical environments are caused by the higher entropy of the native state in water relative to vacuum. The concept of a helical propensity has to be extended to incorporate environmental solvent effects.

Anal paracoccidioidomycosis: an unusual presentation of disseminated disease.

We report a patient with an unusual anal ulceration. The biopsy of an anal lesion and subsequent studies revealed a disseminated form of paracoccidioidomycosis, observed in the lungs, small and large bowel. The anorectal disease frequently represents a secondary site of disease, and the patient must be better evaluated.

Protein-induced membrane disorder: a molecular dynamics study of melittin in a dipalmitoylphosphatidylcholine bilayer.

A molecular dynamics simulation of melittin in a hydrated dipalmitoylphosphatidylcholine (DPPC) bilayer was performed. The 19, 000-atom system included a 72-DPPC phospholipid bilayer, a 26-amino acid peptide, and more than 3000 water molecules. The N-terminus of the peptide was protonated and embedded in the membrane in a transbilayer orientation perpendicular to the surface. The simulation results show that the peptide affects the lower (intracellular) layer of the bilayer more strongly than the upper (extracellular) layer. The simulation results can be interpreted as indicating an increased level of disorder and structural deformation for lower-layer phospholipids in the immediate vicinity of the peptide. This conclusion is supported by the calculated deuterium order parameters, the observed deformation at the intracellular interface, and an increase in fractional free volume. The upper layer was less affected by the embedded peptide, except for an acquired tilt relative to the bilayer normal. The effect of melittin on the surrounding membrane is localized to its immediate vicinity, and its asymmetry with respect to the two layers may result from the fact that it is not fully transmembranal. Melittin's hydrophilic C-terminus anchors it at the extracellular interface, leaving the N-terminus "loose" in the lower layer of the membrane. In general, the simulation supports a role for local deformation and water penetration in melittin-induced lysis. As for the peptide, like other membrane-embedded polypeptides, melittin adopts a significant 25 degree tilt relative to the membrane normal. This tilt is correlated with a comparable tilt of the lipids in the upper membrane layer. The peptide itself retains an overall helical structure throughout the simulation (with the exception of the three N-terminal residues), adopting a 30 degree intrahelical bend angle.

The source of heterogeneity in the heme vicinity of ferricytochrome c.

Heterogeneity in the heme vicinity of ferricytochrome c was reported to be detectable by a split of the NMR signal of the heme methyl 3 group [P.D. Burns and G.N. La Mar, J. Am. Chem. Soc. 101 (1979) 5844]. Using cytochrome c mutants and computer simulations of the native and mutated cytochromes, the source of this heterogeneity is found to originate from the His-33 residue motions. The H33F mutation abolished the NMR split and computer simulations of the H33F mutant revealed a narrower distribution of fluctuations of the radius of gyration, suggesting a more rigid structure due to the mutation. The stabilization of the mutant was further demonstrated by a reduction in the H33F mutant of 4 Kcal/mol in the calculated interaction energy between residue 33 and the rest of the cytochrome, in keeping with known experimental results.

Mechanism of GnRH receptor signaling: from the membrane to the nucleus.

The purpose of this review is to update the information concerning the intracellular effect of GnRH. Binding of GnRH to a G-protein coupled receptor leads to stimulation of Gq and/or G11 protein and to activation of phospholipase C beta. Inositol 1-4-5-triphosphate and early diacylylycerol are the second messengers required for conventional protein kinase C activation. Activation of phospholipase A2 and phospholipase D are also involved, as demonstrated by the liberation of Arachidonic Acid and Phosphatidic Acid. Pituitary cells also express atypical protein kinase C isoforms which mode of activation is not known. Hypothesis concerning transcriptional regulation are presented.

The interaction of borate ions with cytochrome c surface sites: a molecular dynamics study.

Ionic interactions of cytochrome c play an important role in the electron transfer process. Molecular dynamics simulations of the binding of borate ion, which serves as a model ion, at three different cytochrome c surface sites are performed. This work is motivated by previous NMR studies of cytochrome c in borate solution, which indicate the existence of two types of binding sites, a slow exchange site and a fast exchange site. These two types of binding behavior were observed in the dynamic simulations, offering a molecular interpretation of "loose" and "tight" binding. At the "loose" binding sites (near Lys25/Lys27 and Lys55/Lys73) the ion forms two to three hydrogen bonds to the nearest lysine residue. This binding is transient on the time scale of the simulation, demonstrating the feasibility of fast exchange. At the "tight" binding site (near Lys13/Lys86), on the other hand, the ion becomes integrated into the protein hydrogen bond network and remains there for the duration of the simulation (exemplifying slow exchange). Binding simulations of the ion at the "tight" site of H26Q mutant cytochrome c also showed integration of the ion into the protein's hydrogen bond network. However, this integration differs in details from the binding of the ion to the native protein, in agreement with previous NMR observations.

Geometric versus topological clustering: an insight into conformation mapping.

Clustering molecular conformations into "families" is a common procedure in conformational analysis of molecular systems. An implicit assumption which often underlies this clustering approach is that the resulting geometric families reflect the energetic structure of the system's potential energy surface. In a broader context we address the question whether structural similarity is correlated with energy basins, i.e., whether conformations that belong to the same energy basin are also geometrically similar. 'Topological mapping' and principal coordinate projections are used here to address this question and to assess the quality of the 'family clustering' procedure. Applying the analysis to a small tetrapeptide it was found that the general correlation that exists between energy basins and structural similarity is not absolute. Clusters generated by the geometric 'family clustering' procedure do not always reflect the underlying energy basins. In particular it was found that the 'family tree' that is generated by the 'family clustering' procedure is completely inconsistent with its real topological counterpart, the 'disconnectivity' graph of this system. It is also demonstrated that principal coordinate analysis is a powerful visualization technique which, at least for this system, works better when distances are measured in dihedral angle space rather than in cartesian space.