Molecular model for the self-assembly of the cyclic lipodepsipeptide pseudodesmin A

Self-assembly of peptides into supramolecular structures represents an active field of research with potential applications ranging from material science to medicine. Their study typically involves the application of a large toolbox of spectroscopic and imaging techniques. However, quite often, the structural aspects remain underexposed. Besides, molecular modeling of the self-assembly process is usually difficult to handle, since a vast conformational space has to be sampled. Here, we have used an approach that combines short molecular dynamics simulations for peptide dimerization and NMR restraints to build a model of the supramolecular structure from the dimeric units. Experimental NMR data notably provide crucial information about the conformation of the monomeric units, the supramolecular assembly dimensions, and the orientation of the individual peptides within the assembly. This in silico/in vitro mixed approach enables us to define accurate atomistic models of supramolecular structures of the bacterial cyclic lipodepsipeptide pseudodesmin A

A new in-silico method for determination of helical transmembrane domains based on the PepLook scan: application to IL-2Rβ and IL-2Rγc receptor chains

BACKGROUND: Modeling of transmembrane domains (TMDs) requires correct prediction of interfacial residues for in-silico modeling and membrane insertion studies. This implies the defining of a target sequence long enough to contain interfacial residues. However, too long sequences induce artifactual polymorphism: within tested modeling methods, the longer the target sequence, the more variable the secondary structure, as though the procedure were stopped before the end of the calculation (which may in fact be unreachable). Moreover, delimitation of these TMDs can produce variable results with sequence based two-dimensional prediction methods, especially for sequences showing polymorphism. To solve this problem, we developed a new modeling procedure using the PepLook method. We scanned the sequences by modeling peptides from the target sequence with a window of 19 residues. RESULTS: Using sequences whose NMR-structures are already known (GpA, EphA1 and Erb2-HER2), we first determined that the hydrophobic to hydrophilic accessible surface area ratio (ASAr) was the best criterion for delimiting the TMD sequence. The length of the helical structure and the Impala method further supported the determination of the TMD limits. This method was applied to the IL-2Rbeta and IL-2Rgamma TMD sequences of Homo sapiens, Rattus norvegicus, Mus musculus and Bos taurus. CONCLUSIONS: We succeeded in reducing the variation in the TMD limits to only 2 residues and in gaining structural information

Distantly related lipocalins share two conserved clusters of hydrophobic residues: use in homology modeling

BACKGROUND: Lipocalins are widely distributed in nature and are found in bacteria, plants, arthropoda and vertebra. In hematophagous arthropods, they are implicated in the successful accomplishment of the blood meal, interfering with platelet aggregation, blood coagulation and inflammation and in the transmission of disease parasites such as Trypanosoma cruzi and Borrelia burgdorferi. The pairwise sequence identity is low among this family, often below 30%, despite a well conserved tertiary structure. Under the 30% identity threshold, alignment methods do not correctly assign and align proteins. The only safe way to assign a sequence to that family is by experimental determination. However, these procedures are long and costly and cannot always be applied. A way to circumvent the experimental approach is sequence and structure analyze. To further help in that task, the residues implicated in the stabilisation of the lipocalin fold were determined. This was done by analyzing the conserved interactions for ten lipocalins having a maximum pairwise identity of 28% and various functions. RESULTS: It was determined that two hydrophobic clusters of residues are conserved by analysing the ten lipocalin structures and sequences. One cluster is internal to the barrel, involving all strands and the 310 helix. The other is external, involving four strands and the helix lying parallel to the barrel surface. These clusters are also present in RaHBP2, a unusual "outlier" lipocalin from tick Rhipicephalus appendiculatus. This information was used to assess assignment of LIR2 a protein from Ixodes ricinus and to build a 3D model that helps to predict function. FTIR data support the lipocalin fold for this protein. CONCLUSION: By sequence and structural analyzes, two conserved clusters of hydrophobic residues in interactions have been identified in lipocalins. Since the residues implicated are not conserved for function, they should provide the minimal subset necessary to confer the lipocalin fold. This information has been used to assign LIR2 to lipocalins and to investigate its structure/function relationship. This study could be applied to other protein families with low pairwise similarity, such as the structurally related fatty acid binding proteins or avidins.Journal ArticleResearch Support, Non-U.S. Gov'tSCOPUS: ar.jinfo:eu-repo/semantics/publishe

How different serols contribute to cytotoxic action of sea cucumber saponins

Saponins form a diverse class of secondary metabolites found in both plants and some marine invertebrates. Holothuroids, or sea cucumbers, produce these molecules as a chemical defense against predators and parasites, but interestingly, tolerate the cytotoxic nature of these chemicals. This tolerance is poorly understood. The aim of this study was therefore to elucidate the mechanisms behind the tolerance of holothuroid cells to the cytotoxic saponins (e.g. Frondoside A) they produce. This investigation was conducted using a suite of complementary biophysical tools, firstly using in vitro techniques such as Isothermal Titration Calorimetry (ITC) and calcein leakage experiments, for measuring interactions between lipid models of various compositions and the saponin Frondoside A, then using in silico approaches such as docking methods and insertion into implicit lipid bilayers, to provide a molecular point of view on our observations. Interactions between Frondoside A and cholesterol were more favorable than those with the holothuroid Δ7 and Δ9(11) sterols. Liposomes containing cholesterol resulted in exothermic interactions with the holothuroid saponin Frondoside A whereas liposomes containing the Δ7 sterol resulted in endothermic interactions. Lipid phase simulations using settings previously developed for plant saponins revealed that the holothuroid saponin Frondoside A has an agglomerating effect on cholesterol domains, similar to that previously observed for the plant saponin α-Hederin. However, when interacting with the Δ7 sterols, the sterol domains were fragmented into small clusters. A significantly lower leakage was observed with liposomes containing the Δ7 holothuroid sterol than that with liposomes containing cholesterol. Our results suggest that the structural peculiarities of holothuroid sterols provide the organisms with a mechanism to mitigate the sterol-agglomerating effect of saponins on the cell membranes, and therefore to protect sea cucumber cells from the cytotoxicity of the saponins they produce

Use of molecular dynamics simulations to study the interactions between barley allelochemicals and plant plasma membrane

Gramine and hordenine, two alkaloids produced by barley, were shown to inhibit the growth of a common weed (Matricaria recutita L.). This feature could be useful in order to reach a more sustainable weeds management. In vitro experiments have proven that both molecules do interact with lipid bilayers (made of a phosphatidylglycerol (PG) lipid) mimicking plant plasma membranes and are able to modify some of their properties. Moreover, gramine was shown to be more effective than hordenine in both inhibiting weeds growth and altering lipid bilayers properties, suggesting that interactions with membranes could be linked to their mode of action. Molecular dynamics (MD) simulations are carried out in order to get an insight into the molecular mechanisms that underlie these interactions with model membranes and to discriminate between gramine behavior and hordenine behavior

Lipids or Proteins: Who Is Leading the Dance at Membrane Contact Sites?

Understanding the mode of action of membrane contact sites (MCSs) across eukaryotic organisms at the near-atomic level to infer function at the cellular and tissue levels is a challenge scientists are currently facing. These peculiar systems dedicated to inter-organellar communication are perfect examples of cellular processes where the interplay between lipids and proteins is critical. In this mini review, we underline the link between membrane lipid environment, the recruitment of proteins at specialized membrane domains and the function of MCSs. More precisely, we want to give insights on the crucial role of lipids in defining the specificity of plant endoplasmic reticulum (ER)-plasma membrane (PM) MCSs and we further propose approaches to study them at multiple scales. Our goal is not so much to go into detailed description of MCSs, as there are numerous focused reviews on the subject, but rather try to pinpoint the critical elements defining those structures and give an original point of view by considering the subject from a near-atomic angle with a focus on lipids. We review current knowledge as to how lipids can define MCS territories, play a role in the recruitment and function of the MCS-associated proteins and in turn, how the lipid environment can be modified by proteins

Text Line Segmentation of Historical Documents: a Survey

There is a huge amount of historical documents in libraries and in various National Archives that have not been exploited electronically. Although automatic reading of complete pages remains, in most cases, a long-term objective, tasks such as word spotting, text/image alignment, authentication and extraction of specific fields are in use today. For all these tasks, a major step is document segmentation into text lines. Because of the low quality and the complexity of these documents (background noise, artifacts due to aging, interfering lines),automatic text line segmentation remains an open research field. The objective of this paper is to present a survey of existing methods, developed during the last decade, and dedicated to documents of historical interest.Comment: 25 pages, submitted version, To appear in International Journal on Document Analysis and Recognition, On line version available at http://www.springerlink.com/content/k2813176280456k3

Linoleic and linolenic acid hydroperoxides interact differentially with biomimetic plant membranes in a lipid specific manner

Linoleic and linolenic acid hydroperoxides (HPOs) constitute key intermediate oxylipins playing an important role as signaling molecules during plant defense processes in response to biotic or abiotic stress. They have also been demonstrated in vitro as antimicrobial agents against plant fungi and bacteria. To reach the phytopathogens in vivo, the HPOs biosynthesized in the plant cells must cross the plant plasma membrane (PPM) where they can also interact with plasma membrane lipids and have an effect on their organization.In the present study, we have investigated the interaction properties of HPOs with PPM at a molecular level using biophysical tools combining in vitro and in silico approaches and using plant biomimetic lipid systems. Our results have shown that HPOs are able to interact with PPM lipids and perturb their lateral organization. Glucosylceramide (GluCer) is a privileged partner, sitosterol lessens their binding and the presence of both GluCer and sitosterol further reduces their interaction. Hydrophobic effect and polar interactions are involved in the binding. The chemical structure of HPOs influences their affinity for PPM lipids. The presence of three double bonds in the HPO molecule gives rise to a higher affinity comparatively to two double bonds, which can be explained by their differential interaction with the lipid polar headgroups.ARC-FIELD project 13/17-1

Homology modeling and in vivo functional characterization of the zinc permeation pathway in a heavy metal P-type ATPase

The P1B ATPase Heavy Metal ATPase 4 (HMA4) is responsible for zinc and cadmium translocation from roots to shoots in the plant Arabidopsis thaliana. It couples ATP hydrolysis to cytosolic domain movements enabling metal transport across the membrane. Thanks to high conservation level within the P-type ATPase family, the role of the HMA4 cytoplasmic catalytic domains can be inferred from well characterized pumps. In contrast, the function of its terminal cytosolic extensions as well as the metal permeation mechanism through the membrane remains elusive. Here, homology modeling of the HMA4 transmembrane region was conducted based on the crystal structure of a ZntA bacterial homolog. The analysis highlighted amino acids forming a metal permeation pathway, whose importance was subsequently investigated functionally through mutagenesis and complementation experiments in plants. Although the zinc pathway displayed overall conservation among the two proteins, significant differences were observed, especially in the entrance area with altered electronegativity and the presence of a salt bridge/H-bond network. The analysis also newly identified amino acids whose mutation results in total or partial loss of the protein function. In addition, comparison of zinc and cadmium accumulation in shoots of A. thaliana complemented lines revealed a number of HMA4 mutants exhibiting different abilities in zinc and cadmium translocation. These observations could be instrumental to design low cadmium accumulating crops, hence decreasing human cadmium exposure