Mechanistic Insight into the Reactivation of BCAII Enzyme from Denatured and Molten Globule States by Eukaryotic Ribosomes and Domain V rRNAs

In all life forms, decoding of messenger-RNA into polypeptide chain is accomplished by the ribosome. Several protein chaperones are known to bind at the exit of ribosomal tunnel to ensure proper folding of the nascent chain by inhibiting their premature folding in the densely crowded environment of the cell. However, accumulating evidence suggests that ribosome may play a chaperone role in protein folding events in vitro. Ribosome-mediated folding of denatured proteins by prokaryotic ribosomes has been studied extensively. The RNA-assisted chaperone activity of the prokaryotic ribosome has been attributed to the domain V, a span of 23S rRNA at the intersubunit side of the large subunit encompassing the Peptidyl Transferase Centre. Evidently, this functional property of ribosome is unrelated to the nascent chain protein folding at the exit of the ribosomal tunnel. Here, we seek to scrutinize whether this unique function is conserved in a primitive kinetoplastid group of eukaryotic species Leishmania donovani where the ribosome structure possesses distinct additional features and appears markedly different compared to other higher eukaryotic ribosomes. Bovine Carbonic Anhydrase II (BCAII) enzyme was considered as the model protein. Our results manifest that domain V of the large subunit rRNA of Leishmania ribosomes preserves chaperone activity suggesting that ribosome-mediated protein folding is, indeed, a conserved phenomenon. Further, we aimed to investigate the mechanism underpinning the ribosome-assisted protein reactivation process. Interestingly, the surface plasmon resonance binding analyses exhibit that rRNA guides productive folding by directly interacting with molten globule-like states of the protein. In contrast, native protein shows no notable affinity to the rRNA. Thus, our study not only confirms conserved, RNA-mediated chaperoning role of ribosome but also provides crucial insight into the mechanism of the process

Role of MBL2 polymorphisms in sepsis and survival: A pilot study and in silico analysis

Sepsis is a serious infection-induced syndrome with serious ramifications, especially in intensive care units. Global concern motivated the investigation of the role of related genes’ polymorphism in predicting the liability to infection, sepsis, septic shock and survival. Among these genes is the gene encoding mannose-binding lectin (MBL), with its remarkable importance in the immune system. However, the previous studies showed conflicting results and ambiguity that urged us to engage with this issue in the Egyptian population. Prediction of functional and structural impacts of single nucleotide polymorphisms (SNPs) was done using in silico methods. A prospective observational study was conducted in intensive care units; one hundred and thirty patients were followed up. Genotyping was performed using real-time polymerase chain reaction (RT-PCR) technology. MBL SNPs showed a remarkable high frequency in our population, as well. No significant association was found between MBL2 genotypes and any of our analyses (sepsis, septic shock and survival). Only septic shock and age were independently associated with time of survival by Cox regression analysis. Our study may confirm the redundancy of MBL and the absence of significant impact on sepsis liability and mortality in adult patients

Enzootic patterns of Middle East respiratory syndrome coronavirus in imported African and local Arabian dromedary camels: a prospective genomic study

BACKGROUND: The Middle East respiratory syndrome coronavirus (MERS-CoV) is a lethal zoonotic pathogen endemic to the Arabian Peninsula. Dromedary camels are a likely source of infection and the virus probably originated in Africa. We studied the genetic diversity, geographical structure, infection prevalence, and age-associated prevalence among camels at the largest entry port of camels from Africa into the Arabian Peninsula. METHODS: In this prospective genomic study, we took nasal samples from camels imported from Sudan and Djibouti into the Port of Jeddah in Jeddah, Saudi Arabia, over an almost 2-year period and local Arabian camels over 2 months in the year after surveillance of the port. We determined the prevalence of MERS-CoV infection, age-associated patterns of infection, and undertook phylogeographical and migration analyses to determine intercountry virus transmission after local lineage establishment. We compared all virological characteristics between the local and imported cohorts. We compared major gene deletions between African and Arabian strains of the virus. Reproductive numbers were inferred with Bayesian birth death skyline analyses. FINDINGS: Between Aug 10, 2016, and May 3, 2018, we collected samples from 1196 imported camels, of which 868 originated from Sudan and 328 from Djibouti, and between May 1, and June 25, 2018, we collected samples from 472 local camels, of which 189 were from Riyadh and 283 were from Jeddah, Saudi Arabia. Virus prevalence was higher in local camels than in imported camels (224 [47·5%] of 472 vs 157 [13·1%] of 1196; p<0·0001). Infection prevalence peaked among camels older than 1 year and aged up to 2 years in both groups, with 255 (66·9%) of 381 positive cases in this age group. Although the overall geographical distribution of the virus corresponded with the phylogenetic tree topology, some virus exchange was observed between countries corresponding with trade routes in the region. East and west African strains of the virus appear to be geographically separated, with an origin of west African strains in east Africa. African strains of the virus were not re-sampled in Saudi Arabia despite sampling approximately 1 year after importation from Africa. All local Arabian samples contained strains of the virus that belong to a novel recombinant clade (NRC) first detected in 2014 in Saudi Arabia. Reproduction number estimates informed by the sequences suggest sustained endemicity of NRC, with a mean Re of 1·16. INTERPRETATION: Despite frequent imports of MERS-CoV with camels from Africa, African lineages of MERS-CoV do not establish themselves in Saudi Arabia. Arabian strains of the virus should be tested for changes in virulence and transmissibility. FUNDING: German Ministry of Research and Education, EU Horizon 2020, and Emerging Diseases Clinical Trials Partnership

Preliminary fabrication and characterization of electron beam melted Ti-6Al-4V customized dental implant

The current study was aimed to fabricate customized root form dental implant using additive manufacturing technique for the replacement of missing teeth. The root form dental implant was designed using Geomagic (TM) and Magics (TM), the designed implant was directly manufactured by layering technique using ARCAM A2 (TM) electron beam melting system by employing medical grade Ti- 6Al-4V alloy powder. Furthermore, the fabricated implant was characterized in terms of certain clinically important parameters such as surface microstructure, surface topography, chemical purity and internal porosity. Results confirmed that, fabrication of customized dental implants using additive rapid manufacturing technology offers an attractive method to produce extremely pure form of customized titanium dental implants, the rough and porous surface texture obtained is expected to provide better initial implant stabilization and superior osseointegration. (C) 2016 The Authors. Production and hosting by Elsevier B.V. on behalf of King Saud University

Application of Quality of Experience in Networked Services: Review, Trend & Perspectives

Full text embargoed until 17.10.2019 (publisher's embargo period, 12 months

TMX2 Is a Crucial Regulator of Cellular Redox State, and Its Dysfunction Causes Severe Brain Developmental Abnormalities.

The redox state of the neural progenitors regulates physiological processes such as neuronal differentiation and dendritic and axonal growth. The relevance of endoplasmic reticulum (ER)-associated oxidoreductases in these processes is largely unexplored. We describe a severe neurological disorder caused by bi-allelic loss-of-function variants in thioredoxin (TRX)-related transmembrane-2 (TMX2); these variants were detected by exome sequencing in 14 affected individuals from ten unrelated families presenting with congenital microcephaly, cortical polymicrogyria, and other migration disorders. TMX2 encodes one of the five TMX proteins of the protein disulfide isomerase family, hitherto not linked to human developmental brain disease. Our mechanistic studies on protein function show that TMX2 localizes to the ER mitochondria-associated membranes (MAMs), is involved in posttranslational modification and protein folding, and undergoes physical interaction with the MAM-associated and ER folding chaperone calnexin and ER calcium pump SERCA2. These interactions are functionally relevant because TMX2-deficient fibroblasts show decreased mitochondrial respiratory reserve capacity and compensatory increased glycolytic activity. Intriguingly, under basal conditions TMX2 occurs in both reduced and oxidized monomeric form, while it forms a stable dimer under treatment with hydrogen peroxide, recently recognized as a signaling molecule in neural morphogenesis and axonal pathfinding. Exogenous expression of the pathogenic TMX2 variants or of variants with an in vitro mutagenized TRX domain induces a constitutive TMX2 polymerization, mimicking an increased oxidative state. Altogether these data uncover TMX2 as a sensor in the MAM-regulated redox signaling pathway and identify it as a key adaptive regulator of neuronal proliferation, migration, and organization in the developing brain

Estrogen Promotes Mandibular Condylar Fibrocartilage Chondrogenesis and Inhibits Degeneration via Estrogen Receptor Alpha in Female Mice

Temporomandibular joint degenerative disease (TMJ-DD) is a chronic form of TMJ disorder that specifically afflicts people over the age of 40 and targets women at a higher rate than men. Prevalence of TMJ-DD in this population suggests that estrogen loss plays a role in the disease pathogenesis. Thus, the goal of the present study was to determine the role of estrogen on chondrogenesis and homeostasis via estrogen receptor alpha (ERα) during growth and maturity of the joint. Young and mature WT and ERαKO female mice were subjected to ovariectomy procedures and then given placebo or estradiol treatment. The effect of estrogen via ERα on fibrocartilage morphology, matrix production, and protease activity was assessed. In the young mice, estrogen via ERα promoted mandibular condylar fibrocartilage chondrogenesis partly by inhibiting the canonical Wnt signaling pathway through upregulation of sclerostin (Sost). In the mature mice, protease activity was partly inhibited with estrogen treatment via the upregulation and activity of protease inhibitor 15 (Pi15) and alpha-2- macroglobulin (A2m). The results from this work provide a mechanistic understanding of estradiol on TMJ growth and homeostasis and can be utilized for development of therapeutic targets to promote regeneration and inhibit degeneration of the mandibular condylar fibrocartilage.National Institute of Dental & Craniofacial Research of the National Institutes of Health under Award Numbers R56DE020097 (SW) and F32DE026366 (JR

Understanding the Origins of Bacterial Resistance to Aminoglycosides through Molecular Dynamics Mutational Study of the Ribosomal A-Site

Paromomycin is an aminoglycosidic antibiotic that targets the RNA of the bacterial small ribosomal subunit. It binds in the A-site, which is one of the three tRNA binding sites, and affects translational fidelity by stabilizing two adenines (A1492 and A1493) in the flipped-out state. Experiments have shown that various mutations in the A-site result in bacterial resistance to aminoglycosides. In this study, we performed multiple molecular dynamics simulations of the mutated A-site RNA fragment in explicit solvent to analyze changes in the physicochemical features of the A-site that were introduced by substitutions of specific bases. The simulations were conducted for free RNA and in complex with paromomycin. We found that the specific mutations affect the shape and dynamics of the binding cleft as well as significantly alter its electrostatic properties. The most pronounced changes were observed in the U1406C∶U1495A mutant, where important hydrogen bonds between the RNA and paromomycin were disrupted. The present study aims to clarify the underlying physicochemical mechanisms of bacterial resistance to aminoglycosides due to target mutations

A microleakage study of gutta-percha/AH Plus and Resilon/Real self-etch systems after different irrigation protocols

The development and maintenance of the sealing of the root canal system is the key to the success of root canal treatment. The resin-based adhesive material has the potential to reduce the microleakage of the root canal because of its adhesive properties and penetration into dentinal walls. Moreover, the irrigation protocols may have an influence on the adhesiveness of resin-based sealers to root dentin. Objective: The objective of the present study was to evaluate the effect of different irrigant protocols on coronal bacterial microleakage of gutta-percha/AH Plus and Resilon/Real Seal Self-etch systems. Material and Methods: One hundred ninety pre-molars were used. The teeth were divided into 18 experimental groups according to the irrigation protocols and filling materials used. The protocols used were: distilled water; sodium hypochlorite (NaOCl)+eDTA; NaOCl+H3PO4; NaOCl+eDTA+chlorhexidine (CHX); NaOCl+H3PO4+CHX; CHX+eDTA; CHX+ H3PO4; CHX+eDTA+CHX and CHX+H3PO4+CHX. Gutta-percha/AH Plus or Resilon/Real Seal Se were used as root-filling materials. The coronal microleakage was evaluated for 90 days against Enterococcus faecalis. Data were statistically analyzed using Kaplan-Meier survival test, Kruskal-Wallis and Mann-Whitney tests. Results: No significant difference was verified in the groups using chlorhexidine or sodium hypochlorite during the chemo-mechanical preparation followed by eDTA or phosphoric acid for smear layer removal. The same results were found for filling materials. However, the statistical analyses revealed that a final flush with 2% chlorhexidine reduced significantly the coronal microleakage. Conclusion: A final flush with 2% chlorhexidine after smear layer removal reduces coronal microleakage of teeth filled with gutta-percha/AH Plus or Resilon/Real Seal SE