Mapping genotype-phenotype associations of nsSNPs in coiled-coil oligomerization domains of the human proteome.

We assessed the impact of disease mutations (DMs) versus polymorphisms (PYs) in coiled-coil (CC) domains in UniProt by modeling the structural and functional impact of variants in silico with the CC prediction program Multicoil. The structural impact of variants was evaluated with respect to three main metrics: the oligomerization score-to determine whether the variant is stabilizing or destabilizing-the oligomerization state, and the register-specific score. The functional impact was queried indirectly in several ways. First, we examined marginally stable CCs that were either stabilized or destabilized by the variant. Second, we looked for variants that altered the register of the wild-type CC near wild-type irregularities of likely functional importance, such as skips and stammers. Third, we searched for variants that altered the oligomerization state of the CC. DMs tended to be more destabilizing than PYs; but interestingly, PYs were more frequently associated with predicted changes in the oligomerization state. The functional impact was also queried by testing the association of CC variants with multiple phenotypes, that is, pleiotropy. Mutations in CC regions of proteins cause 155 different phenotypes and are more frequently associated with pleiotropy than proteins in general. Importantly, the CC region itself often encodes the pleiotropy.

Impact of organic and mineral inputs onto soil biological and metabolic activities under a long-term rice-wheat cropping system in sub-tropical Indian Inceptisols.

Long-term use of organic and mineral inputs has an overriding impact on soil biological and metabolic activities and crop management. Farm yard manure (FYM), paddy straw (PS) and green manure (GM, Sesbania sesban L.) were used for 24- years old rice (Oyza sativa L.) -wheat (Triticum aestivum L.) cropping system in sub-tropical India to predict whether the screened soil biological and metabolic activities are correlated with system yield. The integrated approaches viz., NPK + FYM, NPK + PS and NPK + GM significantly increased both rice and wheat yield together by 67.5, 44.4 and 55.4%, respectively over control. However, for a few exceptions both soil microbial activity and metabolic activity were remarkably enhanced under integrated treatment NPK + FYM followed by NPK + PS, and NPK + GM, respectively. Among the studied attributes fluorescein diacetate hydrolyzing, dehydrogenase, ß-glucosidase activity (ß-glu) and microbial biomass C (C(mic)) were screened through principal component (PCA) and discriminate analysis (DA) that explained nearly 89% of total variations of the entire data set. Among the four identified attributes, only ß-glu assay value could predict system yield (R2 = 0.65). Further, estimation of ß-glu activity in soil can predict other soil biological properties (R2 = 0.96).

[Studying the effect of heavy metals on tissue protein of an edible fish Cirrhinus mrigala under the influence of pH and water hardness].

The goal of the study was to investigate the influence of pH and water hardness on protein contents of muscle tissues of Cirrhinus mrigala fingerlings by Fourier transform infrared spectroscopy. FT-IR spectra revealed significant differences in absorbance intensities between control and toxic metal-treated muscle tissues, reflecting a change in protein contents due to heavy metals, nickel and chromium, influence at different pH and water hardness. Metal toxicity is also more pronounced at pH 9.0 than in hard water.

Isolation, purification and characterization of naturally derived Crocetin beta-d-glucosyl ester from Crocus sativus L. against breast cancer and its binding chemistry with ER-alpha/HDAC2.

Saffron plant (Crocus sativus L.) is being used as a source of saffron spice and medicine to cure or prevent different types of diseases including cancers. We report the isolation, characterization of bioactive small molecule ([crocetin (ß-d-glucosyl) ester] from the leaf biowastes of saffron plant of Kashmir, India. MTTC assay and Bio-autography aided approach were used to assess anti-oxidant activity and anti-cancer properties of crocin (s) against DPPH free radical and breast cancer cell line respectively. Crocetin beta-d-glucosyl ester restrained proliferation of human breast adeno-carcinoma cell model (MCF-7) without significantly affecting normal cell line (L-6). Further studies involving molecular mechanics generalized born surface area and molecular docking showed that crocetin beta-d-glucosyl ester exhibits strong affinity for estrogen receptor alpha and histone deacetylase 2 (crucial receptors involved in breast cancer signalling) as evidenced by the negative docking score and binding free energy (BFE) values. Therefore, crocetin beta-d-glucosyl ester from Crocus sativus biowastes showed antiproliferative effect possibly by inhibiting estrogen receptor alpha and HDAC2 mediated signalling cascade.

Enhancement of retentivity and selectivity for PAHs in NP-HPLC by high-density immobilization of poly(4-vinylpyridine) as an organic phase on silica.

Poly(4-vinylpyridine)-modified silica with high grafting density have been prepared by a grafting-from (g-from) approach through radical chain-transfer reactions. The widely used silane coupling agent 3-mercaptopropyltrimethoxysilane was used to prepare thiol-terminated silica. Chain-transfer reaction and polymerization of 4-vinylpyridine was carried out using alpha,alpha'-azobisisobutyronitrile as an initiator. Thiol-terminated silica and polymer-modified silica were both characterized qualitatively and quantitatively. The quantification of the organic phase has been done by thermogravimetric analysis and elemental analysis. Thus, the modified silica was used as a packing material and the retention behavior of polycyclic aromatic hydrocarbons (PAHs) was studied in normal-phase high-performance liquid chromatography. Results were compared with those of poly(4-vinylpyridine)-modified silica prepared by a grafting-to (g-to) approach. Commercially available aminopropyl-bonded silica and bare silica columns were also used as reference columns. The column of poly(4-vinylpyridine)-grafted silica prepared by the g-from method, having higher grafting density, provided the better retentivity and selectivity for PAHs compared to the other reference columns.

Molecular structure and vibrational spectra of 3-chloro-4-fluoro benzonitrile by ab initio HF and density functional method.

In this work, the experimental and theoretical spectra of 3-chloro-4-fluoro benzonitrile (3C4FBN) were studied. The Fourier transform infrared and Fourier transform Raman spectra of 3C4FBN were recorded in the solid phase. The optimized geometry was calculated by HF and B3LYP methods with 6-311++G(d,p) basis set. The harmonic-vibrational frequencies, infrared intensities and Raman scattering activities of the title compound were performed at and HF/B3LYP/6-311++G(d,p) level of theories. The scaled theoretical wave number showed very good agreement with the experimental values. The thermodynamic functions of the title compound was also performed at HF/6-31G(d,p) and B3LYP/6-311++G(d,p) level of theories. A detailed interpretation of the infrared and Raman spectra of 3C4FBN was reported. The theoretical spectrograms for FT-IR and FT-Raman spectra of the title molecule have been constructed.

EPR and Structural Characterization of Water-Soluble Mn2+-Doped Si Nanoparticles.

Water-soluble poly(allylamine) Mn2+-doped Si (SiMn) nanoparticles (NPs) were prepared and show promise for biologically related applications. The nanoparticles show both strong photoluminescence and good magnetic resonance contrast imaging. The morphology and average diameter were obtained through transmission electron microscopy (TEM) and high-resolution transmission electron microscopy (HRTEM); spherical crystalline Si NPs with an average diameter of 4.2 ± 0.7 nm were observed. The doping maximum obtained through this process was an average concentration of 0.4 ± 0.3% Mn per mole of Si. The water-soluble SiMn NPs showed a strong photoluminescence with a quantum yield up to 13%. The SiMn NPs had significant T1 contrast with an r1 relaxivity of 11.1 ± 1.5 mM-1 s-1 and r2 relaxivity of 32.7 ± 4.7 mM-1 s-1 where the concentration is in mM of Mn2+. Dextran-coated poly(allylamine) SiMn NPs produced NPs with T1 and T2 contrast with a r1 relaxivity of 27.1 ± 2.8 mM-1 s-1 and r2 relaxivity of 1078.5 ± 1.9 mM-1 s-1. X-band electron paramagnetic resonance spectra are fit with a two-site model demonstrating that there are two types of Mn2+ in these NP's. The fits yield hyperfine splittings (A) of 265 and 238 MHz with significant zero field splitting (D and E terms). This is consistent with Mn in sites of symmetry lower than tetrahedral due to the small size of the NP's.

Antimicrobial activities of a promising glycolipid biosurfactant from a novel marine Staphylococcus saprophyticus SBPS 15.

Biosurfactants have gained a renewed interest in the recent years for their commercial application in diverse research areas. Recent evidences suggest that the antimicrobial activities exhibited by biosurfactants make them promising molecules for the application in the field of therapeutics. Marine microbes are well known for their unique metabolic and functional properties; however, few reports are available till date regarding their biosurfactant production and antimicrobial potential. In an ongoing survey for bioactive microbial metabolites from microbes isolated from diverse ecological niches, a marine Staphylococcus saprophyticus SBPS 15 isolated from the petroleum hydrocarbon contaminated coastal site, Puducherry, India, was identified as a promising biosurfactant producer based on multiple screening methods. This bacterium exhibited growth-dependent biosurfactant production and the recorded yield was 1.345 ± 0.056 g/L (on dry weight basis). The biosurfactant was purified and chemically characterized as a glycolipid with a molecular mass of 606.7 Da, based on TLC, biochemical estimation methods, FT-IR spectrum and MALDI-TOF-MS analysis. Further, the estimated molecular mass was different from the earlier reports on biosurfactants. This new glycolipid biosurfactant exhibited a board range of pH and temperature stability. Furthermore, it revealed a promising antimicrobial activity against many tested human pathogenic bacterial and fungal clinical isolates. Based on these observations, the isolated biosurfactant from the marine S. saprophyticus revealed board physicochemical stabilities and possess excellent antimicrobial activities which proves its significance for possible use in various therapeutic and biomedical applications. To the best of our knowledge, this is the first report of a biosurfactant from the bacterium, S. saprophyticus.

A scientist's guide for submitting data to ZFIN.

The Zebrafish Model Organism Database (ZFIN; zfin.org) serves as the central repository for genetic and genomic data produced using zebrafish (Danio rerio). Data in ZFIN are either manually curated from peer-reviewed publications or submitted directly to ZFIN from various data repositories. Data types currently supported include mutants, transgenic lines, DNA constructs, gene expression, phenotypes, antibodies, morpholinos, TALENs, CRISPRs, disease models, movies, and images. The rapidly changing methods of genomic science have increased the production of data that cannot readily be represented in standard journal publications. These large data sets require web-based presentation. As the central repository for zebrafish research data, it has become increasingly important for ZFIN to provide the zebrafish research community with support for their data sets and guidance on what is required to submit these data to ZFIN. Regardless of their volume, all data that are submitted for inclusion in ZFIN must include a minimum set of information that describes the data. The aim of this chapter is to identify data types that fit into the current ZFIN database and explain how to provide those data in the optimal format for integration. We identify the required and optional data elements, define jargon, and present tools and templates that can help with the acquisition and organization of data as they are being prepared for submission to ZFIN. This information will also appear in the ZFIN wiki, where it will be updated as our services evolve over time.

Membrane Fusion Mediated Targeted Cytosolic Drug Delivery Through scFv Engineered Sendai Viral Envelopes.

Antibody targeted cytoplasmic delivery of drugs is difficult to achieve as antigen-antibody interaction results in the payload being directed to the endosomal compartment. However, Sendai viral envelopes can bring about cytoplasmic delivery due to F-protein mediated membrane fusion. In this study we have generated and fused a recombinant scFv directed to the onco-fetal antigen, the Placental isozyme of Alkaline Phosphatase (PAP) with the trans-membrane and part of the cytoplasmic domain of the Sendai F protein (F(TMC)). Reconstituted virosomes, having both the fusion protein as well as the native F-protein were able to specifically bind and deliver drugs to PAP expressing cells. About 75% of the delivery was cytoplasmic in nature. Hence, this immuno-virosome, which is devoid of the comparatively more toxic HN protein, has the novel ability to combine specific antibody mediated targeting with cytoplasmic delivery. The scFv ensured specific binding to PAP expressing cells, without cross reacting with the other isozymes of alkaline phosphatase. The advantages of cytoplasmic delivery would include reduced degradation and lowered immunogenicity of the payload and carrier. The ubiquitous expression of PAP on a variety of cancers like seminoma, choriocarcinoma, cervical and breast cancers also suggests its potential usefulness in a number of malignancies.

Novel therapeutics for coronary artery disease from genome-wide association study data.

BACKGROUND: Coronary artery disease (CAD), one of the leading causes of death globally, is influenced by both environmental and genetic risk factors. Gene-centric genome-wide association studies (GWAS) involving cases and controls have been remarkably successful in identifying genetic loci contributing to CAD. Modern in silico platforms, such as candidate gene prediction tools, permit a systematic analysis of GWAS data to identify candidate genes for complex diseases like CAD. Subsequent integration of drug-target data from drug databases with the predicted candidate genes can potentially identify novel therapeutics suitable for repositioning towards treatment of CAD. METHODS: Previously, we were able to predict 264 candidate genes and 104 potential therapeutic targets for CAD using Gentrepid (http://www.gentrepid.org), a candidate gene prediction platform with two bioinformatic modules to reanalyze Wellcome Trust Case-Control Consortium GWAS data. In an expanded study, using five bioinformatic modules on the same data, Gentrepid predicted 647 candidate genes and successfully replicated 55% of the candidate genes identified by the more powerful CARDIoGRAMplusC4D consortium meta-analysis. Hence, Gentrepid was capable of enhancing lower quality genotype-phenotype data, using an independent knowledgebase of existing biological data. Here, we used our methodology to integrate drug data from three drug databases: the Therapeutic Target Database, PharmGKB and Drug Bank, with the 647 candidate gene predictions from Gentrepid. We utilized known CAD targets, the scientific literature, existing drug data and the CARDIoGRAMplusC4D meta-analysis study as benchmarks to validate Gentrepid predictions for CAD. RESULTS: Our analysis identified a total of 184 predicted candidate genes as novel therapeutic targets for CAD, and 981 novel therapeutics feasible for repositioning in clinical trials towards treatment of CAD. The benchmarks based on known CAD targets and the scientific literature showed that our results were significant (p < 0.05). CONCLUSIONS: We have demonstrated that available drugs may potentially be repositioned as novel therapeutics for the treatment of CAD. Drug repositioning can save valuable time and money spent on preclinical and phase I clinical studies.

Potential role of glutathione in evolution of thiol-based redox signaling sites in proteins.

Cysteine is susceptible to a variety of modifications by reactive oxygen and nitrogen oxide species, including glutathionylation; and when two cysteines are involved, disulfide formation. Glutathione-cysteine adducts may be removed from proteins by glutaredoxin, whereas disulfides may be reduced by thioredoxin. Glutaredoxin is homologous to the disulfide-reducing thioredoxin and shares similar binding modes of the protein substrate. The evolution of these systems is not well characterized. When a single Cys is present in a protein, conjugation of the redox buffer glutathione may induce conformational changes, resulting in a simple redox switch that effects a signaling cascade. If a second cysteine is introduced into the sequence, the potential for disulfide formation exists. In favorable protein contexts, a bistable redox switch may be formed. Because of glutaredoxin's similarities to thioredoxin, the mutated protein may be immediately exapted into the thioredoxin-dependent redox cycle upon addition of the second cysteine. Here we searched for examples of protein substrates where the number of redox-active cysteine residues has changed throughout evolution. We focused on cross-strand disulfides (CSDs), the most common type of forbidden disulfide. We searched for proteins where the CSD is present, absent and also found as a single cysteine in protein orthologs. Three different proteins were selected for detailed study-CD4, ERO1, and AKT. We created phylogenetic trees, examining when the CSD residues were mutated during protein evolution. We posit that the primordial cysteine is likely to be the cysteine of the CSD which undergoes nucleophilic attack by thioredoxin. Thus, a redox-active disulfide may be introduced into a protein structure by stepwise mutation of two residues in the native sequence to Cys. By extension, evolutionary acquisition of structural disulfides in proteins can potentially occur via transition through a redox-active disulfide state.

No evidence of mutations in the genes for type I and type II 3 beta-hydroxysteroid dehydrogenase (3 beta HSD) in nonclassical 3 beta HSD deficiency.

Nonclassical 3 beta-hydroxysteroid dehydrogenase/delta 5-delta 4-isomerase deficiency (NC3 beta HSDD) has been diagnosed in hyperandrogenic women with an increasing frequency during the last 14 yr. Fifteen menarcheal women with androgen excess syndrome, diagnosed with NC3 beta HSDD previously were restudied, in 12 after discontinuation of glucocorticoid treatment, in 2 patients never treated with glucocorticoids, and in 1 both before and after glucocorticoid therapy. Each of the 15 patients underwent ACTH stimulation testing, in some cases on multiple occasions. Although some (very few) patients seem to have improved with time, others remained the same or got worse. Molecular DNA analysis was also performed in 6 of the patients, using the strategy successfully used to detect point mutations in the type II 3 beta-hydroxysteroid dehydrogenase (3 beta HSD) gene, which are responsible for classical 3 beta HSD deficiency. This strategy consists of the direct sequencing of polymerase chain reaction-amplified DNA fragments corresponding to the complete coding sequence and all intron-exon junctions and to the 5'- and 3'-noncoding region of this gene. We were unable to demonstrate any mutation of the type II 3 beta HSD gene in these 6 patients. To gain additional information about potential mutations, direct sequencing of the type I 3 beta HSD gene was also performed using this same strategy, and no mutations were found. The present study strongly suggests that unlike the salt-losing and nonsalt-losing forms of classical 3 beta HSD deficiency, NC3 beta HSDD is not due to a mutant type II 3 beta HSD enzyme. However, the possibility remains of a mutation(s) in the unsequenced regions of the type II 3 beta HSD gene or elsewhere, such as in a gene for modulatory protein, playing a specific role in the expression of the type II 3 beta HSD gene. On the other hand, knowing the multiple hormonal controls to which 3 beta HSD activity is subject, it cannot be excluded that at least in some cases, NC3 beta HSDD may be an acquired defect, the result of endogenous or environmental factors.

Nonclassical 3 beta-hydroxysteroid dehydrogenase deficiency: a review of our experience with 25 female patients.

OBJECTIVE: To report 15 new menarcheal women affected with nonclassical 3 beta-hydroxysteroid dehydrogenase deficiency (nonclassical 3 beta-HSD) and evaluation of glucocorticoid therapy in treated patients. DESIGN: Diagnosis of these new patients using a standard adrenocorticotropin test. Effects of glucocorticoid therapy on clinical hormonal and sonographic features of each patient are appreciated for periods varying between 4 months and 7 1/2 years. SETTING: All at The New York Hospital-Cornell Medical Center. The Pediatric Endocrinology Ambulatory Service; the Children's Clinical Research Center Core Laboratories; and the Department of Radiology. PATIENTS, PARTICIPANTS: Fifteen menarcheal women (14 to 30 years of age) newly diagnosed and 10 women previously diagnosed were evaluated for symptoms of hyperandrogenism and/or irregular menses. MAIN OUTCOME MEASURE(S): Positive effect of glucocorticoid therapy on signs and symptoms, hormonal levels, and ovarian imaging. RESULTS: Polycystic ovarian syndrome is noted in approximately half the cases. Glucocorticoid treatment greater than 3 months duration results in a reversal of symptoms in most cases.

Transient kinetics of thermodynamic buffering.

The transient response of mitochondrial ATP production towards perturbations was studied by analyzing the trajectories leading from arbitrary initial conditions of the adenine nucleotide pool to the final steady state. These trajectories were calculated from differential equations based on linear relations between flows and thermodynamic forces of the adenylate kinase system including oxidative phosphorylation. The motion of the system along the trajectories consists of two phases: (1) a rapid phase leading from initial states to a common relaxation curve; and (2) a slow phase leading along the relaxation curve to the final steady state. The first phase corresponds to a motion close to the loci of constant adenylic energy charge. In line with this observation is the finding that the energy charge is a constant of motion of the adenylate kinase reaction. The second phase corresponds to a motion along a relaxation curve characterized by minimal Lyapunov exponents in the concentration space of the adenine nucleotides. Thus, both phases of the transient kinetics can be approximated in terms of thermodynamic functions to a high degree of precision. Incubations with isolated rat liver mitochondria were in excellent agreement with the theoretical predictions. In summary, these studies show that the adenylate kinase system not only optimizes the efficiency of oxidative phosphorylation through thermodynamic buffering but, in addition, also deeply influences the transient response of the whole system.

Leptospira interrogans in the genital tract of sheep. Research on ewes and rams experimentally infected with serovar hardjo (hardjobovis).

To verify if Leptospira hardjo can colonize the male and female genital organs of sheep, 9 animals (6 non pregnant ewes and 3 mature rams) were infected with a strain of L. hardjobovis recently recovered from the kidneys of a seropositive ewe. Postinfection controls (bacteriologic, serologic, immunohistochemistry and electron microscopy) failed to disclose the presence of leptospires in the uterus and oviducts, testicles, epididymis, prostate and bulbourethral glands of animals used for the experiment and slaughtered from 37 to 242 postinfection days. All animals showed a renal localization of L. hardjobovis lasting for the entire period of the study (over 8 months). These results emphasize the important role of sheep as maintenance hosts of the serovar.

Leptospira interrogans serovar hardjo in the kidneys and genital tracts of naturally infected sheep.

A bacteriological study was carried out to identify possible renal and/or genital carriers of Leptospira interrogans serovar hardjo. L. hardjo was found at slaughter in the kidneys of three seropositive ewes, but not in uterus or salpinges of these animals.

Opsoclonus myoclonus syndrome: response to plasmapheresis.

An 18-month-old with idiopathic opsomyoclonus, refractory to therapy with ACTH, corticosteroids, and clonazepam received plasmapheresis along with oral corticosteroids and azathioprine. The subject improved dramatically following this treatment. Anticerebellar antibodies were detected from the plasma in this patient and in a two and a half year-old girl with cerebellar ataxia secondary to an adrenal ganglioneuroma.

Identification of novel therapeutics for complex diseases from genome-wide association data.

BACKGROUND: Human genome sequencing has enabled the association of phenotypes with genetic loci, but our ability to effectively translate this data to the clinic has not kept pace. Over the past 60 years, pharmaceutical companies have successfully demonstrated the safety and efficacy of over 1,200 novel therapeutic drugs via costly clinical studies. While this process must continue, better use can be made of the existing valuable data. In silico tools such as candidate gene prediction systems allow rapid identification of disease genes by identifying the most probable candidate genes linked to genetic markers of the disease or phenotype under investigation. Integration of drug-target data with candidate gene prediction systems can identify novel phenotypes which may benefit from current therapeutics. Such a drug repositioning tool can save valuable time and money spent on preclinical studies and phase I clinical trials. METHODS: We previously used Gentrepid (http://www.gentrepid.org) as a platform to predict 1,497 candidate genes for the seven complex diseases considered in the Wellcome Trust Case-Control Consortium genome-wide association study; namely Type 2 Diabetes, Bipolar Disorder, Crohn's Disease, Hypertension, Type 1 Diabetes, Coronary Artery Disease and Rheumatoid Arthritis. Here, we adopted a simple approach to integrate drug data from three publicly available drug databases: the Therapeutic Target Database, the Pharmacogenomics Knowledgebase and DrugBank; with candidate gene predictions from Gentrepid at the systems level. RESULTS: Using the publicly available drug databases as sources of drug-target association data, we identified a total of 428 candidate genes as novel therapeutic targets for the seven phenotypes of interest, and 2,130 drugs feasible for repositioning against the predicted novel targets. CONCLUSIONS: By integrating genetic, bioinformatic and drug data, we have demonstrated that currently available drugs may be repositioned as novel therapeutics for the seven diseases studied here, quickly taking advantage of prior work in pharmaceutics to translate ground-breaking results in genetics to clinical treatments.

Chemical insight into the origin of red and blue photoluminescence arising from freestanding silicon nanocrystals.

Silicon nanocrystals (Si NCs) are attractive functional materials. They are compatible with standard electronics and communications platforms and are biocompatible. Numerous methods have been developed to realize size-controlled Si NC synthesis. While these procedures produce Si NCs that appear identical, their optical responses can differ dramatically. Si NCs prepared using high-temperature methods routinely exhibit photoluminescence agreeing with the effective mass approximation (EMA), while those prepared via solution methods exhibit blue emission that is somewhat independent of particle size. Despite many proposals, a definitive explanation for this difference has been elusive for no less than a decade. This apparent dichotomy brings into question our understanding of Si NC properties and potentially limits the scope of their application. The present contribution takes a substantial step forward toward identifying the origin of the blue emission that is not expected based upon EMA predictions. It describes a detailed comparison of Si NCs obtained from three of the most widely cited procedures as well as the conversion of red-emitting Si NCs to blue emitters upon exposure to nitrogen-containing reagents. Analysis of the evidence is consistent with the hypothesis that the presence of trace nitrogen and oxygen even at the parts per million level in Si NCs gives rise to the blue emission.