Positive and negative effects of the major mammalian messenger ribonucleoprotein p50 on binding of 40 S ribosomal subunits to the initiation codon of beta-globin mRNA

p50, the major core protein bound to mammalian mRNAs, has been reported to stimulate translation at low p50/mRNA ratios and inhibit translation at high p50/mRNA ratios. This study aims to address the molecular mechanisms underlying these phenomena using the in vitro assembly of 48 S preinitiation complexes from fully purified translational components in the presence or absence of p50 as analyzed by the toeprint assay. With limited concentrations of eIF2, eIF3, and eIF4F, p50 (but not pyrimidine tract-binding protein, which was taken for comparison) strongly stimulates formation of the 48 S preinitiation complexes with beta-globin mRNA. This stimulation is observed when just a few molecules of p50 are bound per molecule of the mRNA. When the amount of p50 in solution is increased over some threshold p50/mRNA ratio, a remarkable repression is observed that can still be relieved by adding more eIF2 and eIF4F. At even higher concentrations of p50, the inhibitory effect becomes irreversible. The threshold ratio depends upon the extent of secondary structure of the 5'-untranslated region linked to the beta-globin coding region. Chemical probing has confirmed that the binding of p50 to mRNA involves only the sugar-phosphate backbone of the mRNA leaving nucleotide bases free for interaction with other messenger ribonucleoprotein (mRNP) components. These data are best compatible with the functional role of p50 as a "manager" of mRNA-protein interactions in mammalian mRNPs.     

Parkin-mediated K63-linked polyubiquitination targets misfolded DJ-1 to aggresomes via binding to HDAC6

Sequestration of misfolded proteins into pericentriolar inclusions called aggresomes is a means that cells use to minimize misfolded protein-induced cytotoxicity. However, the molecular mechanism by which misfolded proteins are recruited to aggresomes remains unclear. Mutations in the E3 ligase parkin cause autosomal recessive Parkinson's disease that is devoid of Lewy bodies, which are similar to aggresomes. Here, we report that parkin cooperates with heterodimeric E2 enzyme UbcH13/Uev1a to mediate K63-linked polyubiquitination of misfolded DJ-1. K63-linked polyubiquitination of misfolded DJ-1 serves as a signal for interaction with histone deacetylase 6, an adaptor protein that binds the dynein-dynactin complex. Through this interaction, misfolded DJ-1 is linked to the dynein motor and transported to aggresomes. Furthermore, fibroblasts lacking parkin display deficits in targeting misfolded DJ-1 to aggresomes. Our findings reveal a signaling role for K63-linked polyubiquitination in dynein-mediated transport, identify parkin as a key regulator in the recruitment of misfolded DJ-1 to aggresomes, and have important implications regarding the biogenesis of Lewy bodies.     

Methods of electroencephalographic signal analysis for detection of small hidden changes

The aim of this study was to select and evaluate methods sensitive to reveal small hidden changes in the electroencephalographic (EEG) signal. Two original methods were considered.     

Formation of truncated peptide by‐products via sequence‐specific formyl group transfer from Trp(For) residues to Nα in the course of Boc‐SPPS

(N^In)‐Formyl protective group of tryptophan has been introduced as a base/nucleophile‐labile protective group. It has long been known that a free Nα‐amino group of the peptide can serve as a nucleophile: an irreversible formyl N^In → NH₂ transfer is consistently observed when deformylation is performed last on an otherwise deprotected peptide that possesses free Nα‐amino group. Obviously, this particular side reaction should be expected any time free amino group is exposed to Trp(For), but, at the best of our knowledge, has never been reported in the course of Boc‐SPPS. In the present communication, we describe a set of appropriately designed model experiments that permitted to detect the title side reaction both in solution and in solid‐phase reactions. We observed intermolecular formyl group transfer with a model compound, Trp(For)‐NH₂. Importantly, we also observed this migration on solid support with the rate roughly estimated to be up to 1% of residues per minute. We also observed that the formyl‐group transfer reaction occurred in a sequence‐dependent manner and was suppressed to a non‐detectable level using ‘in situ neutralization’ technique. Because this side reaction is sequence dependent, there might be situations when the rate of the formation of N_α‐formyl termination by‐products is significant. In other cases, the N_α‐For truncated by‐products would not contaminate the final peptide significantly but still could be a source of microheterogeneity. Copyright © 2013 European Peptide Society and John Wiley & Sons, Ltd.     

Bioinformatic characterization of the 4-Toluene Sulfonate Uptake Permease (TSUP) family of transmembrane proteins

The ubiquitous sequence diverse 4-Toluene Sulfonate Uptake Permease (TSUP) family contains few characterized members and is believed to catalyze the transport of several sulfur-based compounds. Prokaryotic members of the TSUP family outnumber the eukaryotic members substantially, and in prokaryotes, but not eukaryotes, extensive lateral gene transfer occurred during family evolution. Despite unequal representation, homologues from the three taxonomic domains of life share well-conserved motifs. We show that the prototypical eight TMS topology arose from an intragenic duplication of a four transmembrane segment (TMS) unit. Possibly, a two TMS α-helical hairpin structure was the precursor of the 4 TMS repeat unit. Genome context analyses confirmed the proposal of a sulfur-based compound transport role for many TSUP homologues, but functional outliers appear to be prevalent as well. Preliminary results suggest that the TSUP family is a member of a large novel superfamily that includes rhodopsins, integral membrane chaperone proteins, transmembrane electron flow carriers and several transporter families. All of these proteins probably arose via the same pathway: 2→4→8 TMSs followed by loss of a TMS either at the N- or C-terminus, depending on the family, to give the more frequent 7 TMS topology.     

Refolding upon force quench and pathways of mechanical and thermal unfolding of ubiquitin

The refolding from stretched initial conformations of ubiquitin (PDB ID: 1ubq) under the quenched force is studied using the C(alpha)-Gō model and the Langevin dynamics. It is shown that the refolding decouples the collapse and folding kinetics. The force-quench refolding-times scale as tau(F) approximately exp(f(q)Deltax(F)/k(B)T), where f(q) is the quench force and Deltax(F) approximately 0.96 nm is the location of the average transition state along the reaction coordinate given by the end-to-end distance. This value is close to Deltax(F) approximately 0.8 nm obtained from the force-clamp experiments. The mechanical and thermal unfolding pathways are studied and compared with the experimental and all-atom simulation results in detail. The sequencing of thermal unfolding was found to be markedly different from the mechanical one. It is found that fixing the N-terminus of ubiquitin changes its mechanical unfolding pathways much more drastically compared to the case when the C-end is anchored. We obtained the distance between the native state and the transition state Deltax(UF) approximately 0.24 nm, which is in reasonable agreement with the experimental data.     

Stabilization of erythrocytes against oxidative and hypotonic stress by tannins isolated from sumac leaves (Rhus typhina L.) and grape seeds (Vitis vinifera L.)

Erythrocytes are constantly exposed to ROS due to their function in the organism. High tension of oxygen, presence of hemoglobin iron and high concentration of polyunsaturated fatty acids in membrane make erythrocytes especially susceptible to oxidative stress. A comparison of the antioxidant activities of polyphenol-rich plant extracts containing hydrolysable tannins from sumac leaves (Rhus typhina L.) and condensed tannins from grape seeds (Vitis vinifera L.) showed that at the 5-50 μg/ml concentration range they reduced to the same extent hemolysis and glutathione, lipid and hemoglobin oxidation induced by erythrocyte treatment with 400 μM ONOO(-) or 1 mM HClO. However, extract (condensed tannins) from grape seeds in comparison with extract (hydrolysable tannins) from sumac leaves stabilized erythrocytes in hypotonic NaCl solutions weakly. Our data indicate that both hydrolysable and condensed tannins significantly decrease the fluidity of the surface of erythrocyte membranes but the effect of hydrolysable ones was more profound. In conclusion, our results indicate that extracts from sumac leaves (hydrolysable tannins) and grape seeds (condensed tannins) are very effective protectors against oxidative damage in erythrocytes.     

Long-Range Correlations and Memory in the Dynamics of Internet Interdomain Routing

Data transfer is one of the main functions of the Internet. The Internet consists of a large number of interconnected subnetworks or domains, known as Autonomous Systems (ASes). Due to privacy and other reasons the information about what route to use to reach devices within other ASes is not readily available to any given AS. The Border Gateway Protocol (BGP) is responsible for discovering and distributing this reachability information to all ASes. Since the topology of the Internet is highly dynamic, all ASes constantly exchange and update this reachability information in small chunks, known as routing control packets or BGP updates. In the view of the quick growth of the Internet there are significant concerns with the scalability of the BGP updates and the efficiency of the BGP routing in general. Motivated by these issues we conduct a systematic time series analysis of BGP update rates. We find that BGP update time series are extremely volatile, exhibit long-term correlations and memory effects, similar to seismic time series, or temperature and stock market price fluctuations. The presented statistical characterization of BGP update dynamics could serve as a basis for validation of existing and developing better models of Internet interdomain routing.     

Radical surgical treatment of the urinary bladder cancer in patients over the age of 60--our experiences

The goal of the paper was the analysis of patients over the age of 60 suffering from the urinary bladder cancer that underwent radical surgical treatment of the urinary bladder and establishing urine derivation. In the 1972-2008 period 2405 patients with the urinary bladder cancer were treated, 296 (12.3%) of whom underwent radical surgical treatment. The average age was between 60 and 80 years--in 207 (70%) patients. In our patients there were 190 patients (91.6%) with transitional cell cancers. According to TNM classification, T3 stage in 92 (44.4%) patients and T2 stage in 85 (41%) patients were predominant in our study. According to histological criteria, the most common stage was G3 stage--in 151 (73%) patients. Radical cystectomy or combined with urethrectomy was performed in 178 (86%) patients. Unfortunately, in 12% of them (T3 and T4 stages) the inner iliac blood vessels were tied off due to a progressive cancer. The outer supravesical urine derivation (Bricker, U-tubing nephrostomy, and ureterocutaneostomy) was done in 163 (78.7%) patients. The inner derivation (Coffey, ureteroileosigmoidostomy, Mainz-Pouch II) was performed in 17 (8.2%) patients and neovesica (Hautmann, Studer) in 24 (11.5%)patients. There were 74 (35.7%) patients with early postoperative complications. Among them the most dominant were the surgical complications--in 28 (13.5%) patients and distant organ complications--in 22 (10.6%) patients. In 75 (36%)patients with negative nodes the survival rate was 55% after five years. In 73 (35%) patients with positive nodes the survival rate was 27% after five years.