Monovalent ions modulate the flux through multiple folding pathways of an RNA pseudoknot

The functions of RNA pseudoknots (PKs), which are minimal tertiary structural motifs and an integral part of several ribozymes and ribonucleoprotein complexes, are determined by their structure, stability and dynamics. Therefore, it is important to elucidate the general principles governing their thermodynamics/folding mechanisms. Here, we combine experiments and simulations to examine the folding/unfolding pathways of the VPK pseudoknot, a variant of the Mouse Mammary Tumor Virus (MMTV) PK involved in ribosomal frameshifting. Fluorescent nucleotide analogs (2-aminopurine and pyrrolocytidine) placed at different stem/loop positions in the PK, and laser temperature-jump approaches serve as local probes allowing us to monitor the order of assembly of VPK with two helices with different intrinsic stabilities. The experiments and molecular simulations show that at 50 mM KCl the dominant folding pathway populates only the more stable partially folded hairpin. As the salt concentration is increased a parallel folding pathway emerges, involving the less stable hairpin structure as an alternate intermediate. Notably, the flux between the pathways is modulated by the ionic strength. The findings support the principle that the order of PK structure formation is determined by the relative stabilities of the hairpins, which can be altered by sequence variations or salt concentrations. Our study not only unambiguously demonstrates that PK folds by parallel pathways, but also establishes that quantitative description of RNA self-assembly requires a synergistic combination of experiments and simulations.Comment: Supporting Information include

Evaluation of deficit irrigation regime, row spacing and dual plantation of drip irrigated tomato under high tunnel

The present study was carried out at Water Management Research Centre, University of Agriculture, Faisalabad, Pakistan, to investigate the effect of row spacing, deficit irrigation and dual plantation on tomato yield grown in high tunnel under polyethylene black mulch and drip irrigation system. The field experiment layout comprised of three rows spacing (S1, S2 and S3) as (0.45, 0.60, and 0.75 m respectively) and two irrigation levels (I1 and I2) as (100% and 75% of required irrigation respectively). In addition, a dual plantation treatment spaced at 0.45 m was also investigated under both irrigation levels. Results showed that the total fruit yield was significantly influenced by row spacing and irrigation level, however, their interaction was non-significant. The 100% irrigation gave 6.53, 4.49, and 5.94% more yield than 75% irrigation treatment under 0.45, 0.60, and 0.75 m row spacing, respectively. However, the irrigation water use efficiency was found to be higher in deficit treatment (75% irrigation) by 25.16, 27.60, and 25.86% than full irrigation treatments under 0.45, 0.60, and 0.75 m row spacing, respectively. The results of single and dual plantation showed that dual plantation increased the fruit yield by 7.62 and 11.28% than the single plantation under full and deficit irrigation respectively and covering approximately the same area

Genetic Characterization of Carbapenem Resistant Acinetobacter baumannii in Tertiary care settings of Lahore, Pakistan

Background: Acinetobacter baumannii is major cause of ventilator associated pneumoniae (VAP) as it is an opportunistic nosocomial organism. The current study was to find out the antibiotic resistance pattern of Acinetobacter baumannii, its phenotype and the genetic characterization of Metallo-β-Lactamase (MBL) genes that are responsible for carbapenem resistance.Methods: One hundred and fifty Carbapenem resistant Acinetobacter baumannii (CRAB) specimens were isolated and PCR amplification of organism specific bla-OXA-51gene was performed and antibiotic susceptibility was checked. Phenotypic susceptibility analysis was performed by Modified Hodge Test (MHT) and Imipenem-EDTA Double Disc Synergy Test (IMP-EDTA DDST). The carbapenemases and MBL producing genes were amplified by PCR.Results: CRAB showed high resistance against piperacillin/tazobactam (99.3%), cefepime and ceftazidime (99.3% each), amikacin (91.3%), ciprofloxacin (96.7%) and levofloxacin (96.7%). Only one isolate showed resistance to colistin. The isolates positive for both MHT and DDST (n=70) were further characterized to detect metallo-β-lactamase genes. Molecular characterization revealed the presence of bla-OXA-51 gene in all tested isolates (100%) followed by bla-VIM 89%, bla-OXA-23 64%, respectively and so on. Few genes coexisted with each other including bla VIM, bla OXA 23, bla OXA 51 and bla NDM-1.  None of the isolate was found positive for bla-IMP gene.Conclusion: It is concluded that CRAB isolates exhibited a high rate of resistance towards antimicrobials because of the presence of drug hydrolyzing enzymes, carbapenemases and MBLs. This is among the rare study reported recently indicating CRAB isolates co-harboring many resistant genes are very difficult to treat. There is a dire need to develop novel antibiotics against resistant A. baumannii to minimize its prevalence. Moreover, it is recommended that colistin treatment in the clinical settings should be continuously monitored in order to prevent the development of resistance

Loop dependence of the stability and dynamics of nucleic acid hairpins

Hairpin loops are critical to the formation of nucleic acid secondary structure, and to their function. Previous studies revealed a steep dependence of single-stranded DNA (ssDNA) hairpin stability with length of the loop (L) as ∼L8.5 ± 0.5, in 100 mM NaCl, which was attributed to intraloop stacking interactions. In this article, the loop-size dependence of RNA hairpin stabilities and their folding/unfolding kinetics were monitored with laser temperature-jump spectroscopy. Our results suggest that similar mechanisms stabilize small ssDNA and RNA loops, and show that salt contributes significantly to the dependence of hairpin stability on loop size. In 2.5 mM MgCl2, the stabilities of both ssDNA and RNA hairpins scale as ∼L4 ± 0.5, indicating that the intraloop interactions are weaker in the presence of Mg2+. Interestingly, the folding times for ssDNA hairpins (in 100 mM NaCl) and RNA hairpins (in 2.5 mM MgCl2) are similar despite differences in the salt conditions and the stem sequence, and increase similarly with loop size, ∼L2.2 ± 0.5 and ∼L2.6 ± 0.5, respectively. These results suggest that hairpins with small loops may be specifically stabilized by interactions of the Na+ ions with the loops. The results also reinforce the idea that folding times are dominated by an entropic search for the correct nucleating conformation

Serum Alpha 1 Antitrypsin and Pulmonary Emphysema

Using isoelectric focusing (lEE) and radial immunodiffusion (RID) techniques, serum samples from 100 normal healthy adults and 21 patients with pulmonary emphysema were analysed to identify varioud alpha 1 antitrypsin phenotypes and the Seru m concentrations,Ten percent of the patients had low serum values. The normal or most common genetic form, MM, is the predominant phenotype in both controls and patients (JPMA 46:102, 1996)

Evidence for a bind-then-bend mechanism for architectural DNA binding protein yNhp6A

The yeast Nhp6A protein (yNhp6A) is a member of the eukaryotic HMGB family of chromatin factors that enhance apparent DNA flexibility. yNhp6A binds DNA nonspecifically with nM affinity, sharply bending DNA by \u3e60◦. It is not known whether the protein binds to unbent DNA and then deforms it, or if bent DNA conformations are ‘captured’ by protein binding. The former mechanism would be supported by discovery of conditions where unbent DNA is bound by yNhp6A. Here, we employed an array of conformational probes (FRET, fluorescence anisotropy, and circular dichroism) to reveal solution conditions in which an 18- base-pair DNA oligomer indeed remains bound to yNhp6A while unbent. In 100 mM NaCl, yNhp6Abound DNA unbends as the temperature is raised, with no significant dissociation of the complex detected up to ∼45◦C. In 200 mM NaCl, DNA unbending in the intact yNhp6A complex is again detected up to ∼35◦C. Microseconds-resolved laser temperaturejump perturbation of the yNhp6a–DNA complex revealed relaxation kinetics that yielded unimolecular DNA bending/unbending rates on timescales of 500 s−1 ms. These data provide the first direct observation of bending/unbending dynamics of DNA in complex with yNhp6A, suggesting a bind-then-bend mechanism for this protein

An open label hepatoprotective activity of Jawārish bisbāsā in central obesity patients

Liver plays a major role in metabolism and excretion of xenobiotics. Liver dysfunction is a major health problem that challenges health care professionals, pharmaceutical industry and drug regulatory agencies. The most common symptoms are fatigue and discomfort in abdomen while in patients who are obese with BMI > 25, about one third have metabolic syndromes. The present clinical research was conducted in Regional Research Institute of Unani Medicine, Aligarh. The patients under trial were selected from GOPD of institute for obesity and it was observed that out of total registered patients’ approx 28% have elevated liver enzymes from its normal range without any sign and symptom of hepatitis etc. Total 23 patients were selected from the study whose liver enzymes (SGOT, SGPT and ALP) were high at the baseline. Unani pharmacopoeial drug Jawārish bisbāsā 7 g daily was prescribed with lukewarm water in the morning and evening empty stomach for 8 weeks. At the end of study the result were compared with base line. It was observed that Jawārish bisbāsā significantly reduced the liver enzymes, e.g. SGOT, SGPT, Alkaline Phosphatase in comparison to baseline values. It may be concluded that Jawārish bisbāsā showed hepatoprotective effect in this clinical study.

Situational factors shape moral judgements in the trolley dilemma in Eastern, Southern and Western countries in a culturally diverse sample

The study of moral judgements often centres on moral dilemmas in which options consistent with deontological perspectives (that is, emphasizing rules, individual rights and duties) are in conflict with options consistent with utilitarian judgements (that is, following the greater good based on consequences). Greene et al. (2009) showed that psychological and situational factors (for example, the intent of the agent or the presence of physical contact between the agent and the victim) can play an important role in moral dilemma judgements (for example, the trolley problem). Our knowledge is limited concerning both the universality of these effects outside the United States and the impact of culture on the situational and psychological factors affecting moral judgements. Thus, we empirically tested the universality of the effects of intent and personal force on moral dilemma judgements by replicating the experiments of Greene et al. in 45 countries from all inhabited continents. We found that personal force and its interaction with intention exert influence on moral judgements in the US and Western cultural clusters, replicating and expanding the original findings. Moreover, the personal force effect was present in all cultural clusters, suggesting it is culturally universal. The evidence for the cultural universality of the interaction effect was inconclusive in the Eastern and Southern cultural clusters (depending on exclusion criteria). We found no strong association between collectivism/individualism and moral dilemma judgements

Role of Proinsulin Self-Association in Mutant INS Gene–Induced Diabetes of Youth

Abnormal interactions between misfolded mutant and wild-type (WT) proinsulin (PI) in the endoplasmic reticulum (ER) drive the molecular pathogenesis of mutant INS gene-induced diabetes of youth (MIDY). How these abnormal interactions are initiated remains unknown. Normally, PI-WT dimerizes in the ER. Here, we suggest that the normal PI-PI contact surface, involving the B-chain, contributes to dominant-negative effects of misfolded MIDY mutants. Specifically, we find that PI B-chain tyrosine-16 (Tyr-B16), which is a key residue in normal PI dimerization, helps confer dominant-negative behavior of MIDY mutant PI-C(A7)Y. Substitutions of Tyr-B16 with either Ala, Asp, or Pro in PI-C(A7)Y decrease the abnormal interactions between the MIDY mutant and PI-WT, rescuing PI-WT export, limiting ER stress, and increasing insulin production in β-cells and human islets. This study reveals the first evidence indicating that noncovalent PI-PI contact initiates dominant-negative behavior of misfolded PI, pointing to a novel therapeutic target to enhance PI-WT export and increase insulin production