Mesoscopic dynamics of colloids simulated with dissipative particle dynamics and fluid particle model

We report results of numerical simulations of complex fluids, using a combination of discrete-particle methods. Our molecular modeling repertoire comprises three simulation techniques: molecular dynamics (MD), dissipative particle dynamics (DPD), and the fluid particle model (FPM). This type of model can depict multi-resolution molecular structures (see the Figure) found in complex fluids ranging from single micelle, colloidal crystals, large-scale colloidal aggregates up to the mesoscale processes of hydrodynamical instabilities in the bulk of colloidal suspensions. We can simulate different colloidal structures in which the colloidal beds are of comparable size to the solvent particles. This undertaking is accomplished with a two-level discrete particle model consisting of the MD paradigm with a Lennard-Jones (L-J) type potential for defining the colloidal particle system and DPD or FPM for modeling the solvent. We observe the spontaneous emergence of spherical or rod-like micelles and their crystallization in stable hexagonal or worm-like structures, respectively. The ordered arrays obtained by using the particle model are similar to the 2D colloidal crystals observed in laboratory experiments. The micelle shape and its hydrophobic or hydrophilic character depend on the ratio between the scaling factors of the interactions between colloid–colloid to colloid–solvent. Unlike the miscellar arrays, the colloidal aggregates involve the colloid–solvent interactions prescribed by the DPD forces. Different from the assumption of equilibrium growth, the two-level particle model can display much more realistic molecular physics, which allows for the simulation of aggregation for various types of colloids and solvent liquids over a very broad range of conditions. We discuss the potential prospects of combining MD, DPD, and FPM techniques in a single three-level model. Finally, we present results from large-scale simulation of the Rayleigh–Taylor instability and dispersion of colloidal slab in 2D and 3D. Electronic supplementary material to this paper can be obtained by using the Springer LINK server located at http://dx.doi.org/10.1007/s00894-001-0068-3.Electronic Supplementary Material available.     

Hyperresistance to photosensitized lipid peroxidation and apoptotic killing in 5-aminolevulinate-treated tumor cells overexpressing mitochondrial GPX4

Antitumor photodynamic therapy (PDT) with administered 5-aminolevulinic acid (ALA) is based on metabolism of ALA to protoporphyrin IX (PpIX), which acts as a sensitizer of photo-oxidative damage leading to apoptotic or necrotic cell death. An initial goal of this study was to ascertain how the PpIX-sensitized death mechanism for a breast tumor line (COH-BR1 cells) might be influenced by the conditions of ALA exposure in vitro. Two different treatment protocols were developed for addressing this question: (i) continuous incubation with 1 mM ALA for 90 min; and, (ii) discontinuous incubation, i.e., 15 min with 1 mM ALA followed by 225 min without it. Following exposure to 2 J/cm2 of visible light, cell viability, death mechanism, and lipid hydroperoxide (LOOH) level were evaluated for each protocol using thiazolyl blue, Hoechst staining, and HPLC with electrochemical detection assays, respectively. PpIX was found to sensitize apoptosis when it existed mainly in mitochondria (protocol-1), but necrosis when it diffused to other sites, including plasma membrane (protocol-2). Experiments with a transfectant clone, 7G4, exhibiting approximately 85 times greater activity of the LOOH-detoxifying selenoenzyme GPX4 than parental cells, provided additional information about death mechanism. Located predominantly in mitochondria of 7G4 cells, GPX4 strongly inhibited both LOOH accumulation and apoptosis under protocol-1 conditions, but had no significant effect under protocol-2 conditions. These findings support the hypothesis that LOOHs produced by attack of photogenerated singlet oxygen on mitochondrial membrane lipids play an important early role in the apoptotic death cascade.     

Identification and molecular cloning of a gene encoding a fibronectin-binding protein (CadF) from Campylobacter jejuni

Campylobacter jejuni , a Gram-negative bacterium, is a common cause of gastrointestinal disease. By analogy with other enteric pathogens such as Salmonella and Shigella , the ability of C. jejuni to bind to host cells is thought to be essential in the pathogenesis of enteritis. Scanning electron microscopy of infected INT407 cells suggested that C. jejuni bound to a component of the extracellular matrix. Binding assays using immobilized extracellular matrix proteins and soluble fibronectin showed specific and saturable binding of fibronectin to C. jejuni . Ligand immunoblot assays using ¹²⁵I-labelled fibronectin revealed specific binding to an outer membrane protein with an apparent molecular mass of 37 kDa. A rabbit antiserum, raised against the gel-purified protein, reacted with a 37 kDa protein in all C. jejuni isolates ( n  = 15) as tested by immunoblot analysis. Antibodies present in convalescent serum from C. jejuni -infected individuals also recognized a 37 kDa protein. The gene encoding the immunoreactive 37 kDa protein was cloned and sequenced. Sequencing of overlapping DNA fragments revealed an open reading frame (ORF) that encodes a protein of 326 amino acids with a calculated molecular mass of 36 872 Da. The deduced amino acid sequence of the ORF exhibited 52% similarity and 28% identity to the root adhesin protein from Pseudomonas fluorescens . Isogenic C. jejuni mutants which lack the 37 kDa outer membrane protein, which we have termed CadF, displayed significantly reduced binding to fibronectin. Biotinylated fibronectin bound to a protein with an apparent molecular mass of 37 kDa in the outer membrane protein extracts from wild-type C. jejuni as judged by ligand-binding blots. These results indicate that the binding of C. jejuni to fibronectin is mediated by the 37 kDa outer membrane protein which is conserved among C. jejuni isolates.     

Fast assessment of structural models of ion channels based on their predicted current–voltage characteristics

Computational prediction of protein structures is a difficult task, which involves fast and accurate evaluation of candidate model structures. We propose to enhance single‐model quality assessment with a functionality evaluation phase for proteins whose quantitative functional characteristics are known. In particular, this idea can be applied to evaluation of structural models of ion channels, whose main function ‐ conducting ions ‐ can be quantitatively measured with the patch‐clamp technique providing the current–voltage characteristics. The study was performed on a set of KcsA channel models obtained from complete and incomplete contact maps. A fast continuous electrodiffusion model was used for calculating the current–voltage characteristics of structural models. We found that the computed charge selectivity and total current were sensitive to structural and electrostatic quality of models. In practical terms, we show that evaluating predicted conductance values is an appropriate method to eliminate models with an occluded pore or with multiple erroneously created pores. Moreover, filtering models on the basis of their predicted charge selectivity results in a substantial enrichment of the candidate set in highly accurate models. Tests on three other ion channels indicate that, in addition to being a proof of the concept, our function‐oriented single‐model quality assessment method can be directly applied to evaluation of structural models of some classes of protein channels. Finally, our work raises an important question whether a computational validation of functionality should be included in the evaluation process of structural models, whenever possible. Proteins 2016; 84:217–231. © 2015 Wiley Periodicals, Inc.     

Phylogeography of Scots pine in Europe and Asia based on mtDNA polymorphisms

We analyzed mitochondrial DNA polymorphisms to search for evidence of the genetic structure and patterns of admixture in 124 populations (N = 1407 trees) across the distribution of Scots pine in Europe and Asia. The markers revealed only a weak population structure in Central and Eastern Europe and suggested postglacial expansion to middle and northern latitudes from multiple sources. Major mitotype variants include the remnants of Scots pine at the north-western extreme of the distribution in the Scottish Highlands; two main variants (western and central European) that contributed to the contemporary populations in Norway and Sweden; the central-eastern European variant present in the Balkan region, Finland, and Russian Karelia; and a separate one common to most eastern European parts of Russia and western Siberia. We also observe signatures of a distinct refugium located in the northern parts of the Black Sea basin that contributed to the patterns of genetic variation observed in several populations in the Balkans, Ukraine, and western Russia. Some common haplotypes of putative ancient origin were shared among distant populations from Europe and Asia, including the most southern refugial stands that did not participate in postglacial recolonization of northern latitudes. The study indicates different genetic lineages of the species in Europe and provides a set of genetic markers for its finer-scale population history and divergence inference.     

Faithful and efficient translation of viral and cellular eukaryotic mRNAs in a cell-free S-27 extract of Saccharomycescerevisiae

The preparation of yeast spheroplast 27,000 × g supernatant /S-27/ which initiates translation of endogenous and exogenous mRNAs is described. The activity of this protein synthesis system is comparable to that of the most efficient cell-free extracts currently in use. The yeast S-27 system is able to carry out faithful translation of distinct eukaryotic mRNAs into proteins as large as 180,000 daltons.     

Characterization of the U3 and U6 snRNA genes from wheat: U3 snRNA genes in monocot plants are transcribed by RNa polymerase III

We have demonstrated recently that the genes encoding the U3 small nuclear RNA (snRNA) in dicot plants are transcribed by RNA polymerase III (pol III), and not RNA polymerase II (pol II) as in all other organisms studied to date. The U3 gene was the first example of a gene transcribed by different polymerases in different organisms. Based on phylogenetic arguments we proposed that a polymerase specificity change of the U3 snRNA gene promoter occurred during plant evolution. To map such an event we are examining the U3 gene polymerase specificity in other plant species. We report here the characterization of a U3 gene from wheat, a monocot plant. This gene contains the conserved promoter elements, USE and TATA, in a pol III-specific spacing seen also in a wheat U6 snRNA gene characterized in this report. Both the U3 and the U6 genes possess typical pol III termination signals but lack the cis element, responsible for 3-end formation, found in all plant pol II-specific snRNA genes. In addition, expression of the U3 gene in transfected maize protoplasts is less sensitive to -amanitin than a pol II-transcribed U2 gene. Based on these data we conclude that the wheat U3 gene is transcribed by pol III. This observation suggests that the postulated RNA polymerase specificity switch of the U3 gene took place prior to the divergence of angiosperm plants into monocots and dicots.     

Purification and characterization of a chitosanase from Streptomyces N174

A highly efficient chitosanase producer, the actinomycete N174, identified by chemotaxonomic methods as belonging to the genus Streptomyces was isolated from soil. Chitosanase production by N174 was inducible by chitosan or d-glucosamine. In culture filtrates the chitosanase accounted for 50–60% of total extracellular proteins. The chitosanase was purified by polyacrylic acid precipitation, CM-Sepharose and gel permeation chromatography. The maximum velocity of chitosan degradation was obtained at 65° C when the pH was maintained at 5.5. The enzyme degraded chitosans with a range of acetylation degrees from 1 to 60% but not chitin or CM-cellulose. The enzyme showed an endo-splitting type of activity and the end-product of chitosan degradation contained a mixture of dimers and trimers of d-glucosamine.Correspondence to: R. Brzezinski     

Adenomas – Genetic factors in colorectal cancer prevention

Colorectal cancer is the second most common type of cancer both in Europe and Poland. During the last 30 years more than a 3-fold increase has been observed in Poland due to environmental and genetic factors. Almost all colorectal malignancies are related to the formation and malignant transformation of colorectal dysplasia and adenoma. Efforts aiming to decrease the number of colorectal cancer deaths are focused on the disease early detection. Genetic diagnosis for hereditary syndromes predisposing to colorectal cancer has been developed and is a part of the routine treatment. Most cancers are sporadic. They often develop from polyps in the colon. In addition to the genetic events described in the 1990s, showing the adenoma transformation into carcinoma that has been a prime example of malignant transformation for a long time, there are also other possibilities of neoplastic transformation. The recognition of colorectal cancer risk factors make sense as their nature is lifestyle- and diet-related. In this review paper those risk factors are presented and the prevention of colorectal cancer is discussed taking into account genetic factors.     

Plasmid-mediated control of nodulation in Rhizobium trifolii

The nodulation ability was effectively eliminated from different Rhizobium trifolii strains incubated at elevated temperature (urkowski and Lorkiewicz, 1978). Non-nodulating (Nod^-) mutants were stable and no reversion of Nod^- to Nod⁺ phenotype was observed. Strains R. trifolii 24 and T12 which showed a high percentage of elimination of nodulation ability were examined in detail. Two plasmids were detected in strain 24 using neutral and alkaline sucrose gradient centrifugation of plasmid preparations. Molecular weights of the plasmids pWZ1 and pWZ2 were 460 Mdal and 190 Mdal, respectively. Rhizobium lysates labeled with ³H-thymidine and ultracentrifuged in caesium chloride — ethidium bromide gradients demonstrated a 40% reduction of the plasmid DNA content in R. trifolii 24 Nod^- mutants in comparison with the nodulating wild type strain 24. It was found further that non-nodulation of mutants 24 Nod^- was due to the absence of plasmid pWZ2. Sucrose gradient data also demonstrated that strain T12 contained two plasmids with molecular weights corresponding to those of pWZ1 and pWZ2, respectively. In Nod^- mutant clones derived from strain T12, pWZ2 plasmid was missing.Non Standard Abbreviations CCC covalently closed circular - OC open cirucular - Sarkosyl sodium N-lauroylsarcosinate