Hydrogen bond networks determine emergent mechanical and thermodynamic properties across a protein family

<p>Abstract</p> <p>Background</p> <p>Gram-negative bacteria use periplasmic-binding proteins (bPBP) to transport nutrients through the periplasm. Despite immense diversity within the recognized substrates, all members of the family share a common fold that includes two domains that are separated by a conserved hinge. The hinge allows the protein to cycle between open (apo) and closed (ligated) conformations. Conformational changes within the proteins depend on a complex interplay of mechanical and thermodynamic response, which is manifested as an increase in thermal stability and decrease of flexibility upon ligand binding.</p> <p>Results</p> <p>We use a distance constraint model (DCM) to quantify the give and take between thermodynamic stability and mechanical flexibility across the bPBP family. Quantitative stability/flexibility relationships (QSFR) are readily evaluated because the DCM links mechanical and thermodynamic properties. We have previously demonstrated that QSFR is moderately conserved across a mesophilic/thermophilic RNase H pair, whereas the observed variance indicated that different enthalpy-entropy mechanisms allow similar mechanical response at their respective melting temperatures. Our predictions of heat capacity and free energy show marked diversity across the bPBP family. While backbone flexibility metrics are mostly conserved, cooperativity correlation (long-range couplings) also demonstrate considerable amount of variation. Upon ligand removal, heat capacity, melting point, and mechanical rigidity are, as expected, lowered. Nevertheless, significant differences are found in molecular cooperativity correlations that can be explained by the detailed nature of the hydrogen bond network.</p> <p>Conclusion</p> <p>Non-trivial mechanical and thermodynamic variation across the family is explained by differences within the underlying H-bond networks. The mechanism is simple; variation within the H-bond networks result in altered mechanical linkage properties that directly affect intrinsic flexibility. Moreover, varying numbers of H-bonds and their strengths control the likelihood for energetic fluctuations as H-bonds break and reform, thus directly affecting thermodynamic properties. Consequently, these results demonstrate how unexpected large differences, especially within cooperativity correlation, emerge from subtle differences within the underlying H-bond network. This inference is consistent with well-known results that show allosteric response within a family generally varies significantly. Identifying the hydrogen bond network as a critical determining factor for these large variances may lead to new methods that can predict such effects.</p

An unusual case of gout in the wrist: the importance of monitoring medication dosage and interaction. A case report

<p>Abstract</p> <p>Background</p> <p>Gouty arthritis of the wrist is uncommon although gout itself is the most common inflammatory arthritis in older patients. Some known risk factors for the development of gout include trauma, alcohol use, obesity, hyperuricaemia, hypertension and diabetes mellitus. As well, certain medications have been shown to promote the development of gout. These include thiazide diuretics, low dose salicylates and cyclosporine. We present a case of gouty wrist pain possibly precipitated by a medication dosage increase as well as medication interactions.</p> <p>Case presentation</p> <p>A 77 year old male presented with right wrist pain. Redness and swelling was present at the dorsal aspect of his wrist and range of motion was full with pain at end range upon examination. One week prior, his anti-hypertensive medication dosage had been increased. The patient's situation continued to worsen. Radiographic examination revealed changes consistent with gouty arthritis.</p> <p>Conclusion</p> <p>It is important for clinicians treating joint conditions to be aware of patients' comorbidities, medication usage and changes in dosages. Education of patients with gout is of prime importance. Clinicians should educate patients that gout may occur at any joint in the body not only the lower limb. Patients should be aware of the signs and symptoms of an acute gouty attack and be made aware that changes in certain medication dosages may precipitate an attack. Awareness of radiographic changes associated with gout is still of importance although these changes are not seen as frequently as they have been in the past due to better control of the disease.</p

The challenges of communicating research evidence in practice: perspectives from UK health visitors and practice nurses

&lt;p&gt;Background: Health practitioners play a pivotal role in providing patients with up-to-date evidence and health information. Evidence-based practice and patient-centred care are transforming the delivery of healthcare in the UK. Health practitioners are increasingly balancing the need to provide evidence-based information against that of facilitating patient choice, which may not always concur with the evidence base. There is limited research exploring how health practitioners working in the UK, and particularly those more autonomous practitioners such as health visitors and practice nurses working in community practice settings, negotiate this challenge. This research provides a descriptive account of how health visitors and practice nurses negotiate the challenges of communicating health information and research evidence in practice.&lt;/p&gt; &lt;p&gt;Methods: A total of eighteen in-depth telephone interviews were conducted in the UK between September 2008 and May 2009. The participants comprised nine health visitors and nine practice nurses, recruited via adverts on a nursing website, posters at a practitioner conference and through recommendation. Thematic analysis, with a focus on constant comparative method, was used to analyse the data.&lt;/p&gt; &lt;p&gt;Results: The data were grouped into three main themes: communicating evidence to the critically-minded patient; confidence in communicating evidence; and maintaining the integrity of the patient-practitioner relationship. These findings highlight some of the daily challenges that health visitors and practice nurses face with regard to the complex and dynamic nature of evidence and the changing attitudes and expectations of patients. The findings also highlight the tensions that exist between differing philosophies of evidence-based practice and patient-centred care, which can make communicating about evidence a daunting task.&lt;/p&gt; &lt;p&gt;Conclusions: If health practitioners are to be effective at communicating research evidence, we suggest that more research and resources need to be focused on contextual factors, such as how research evidence is negotiated, appraised and communicated within the dynamic patient-practitioner relationship.&lt;/p&gt

Changes in Lysozyme Flexibility upon Mutation Are Frequent, Large and Long-Ranged

We investigate changes in human c-type lysozyme flexibility upon mutation via a Distance Constraint Model, which gives a statistical mechanical treatment of network rigidity. Specifically, two dynamical metrics are tracked. Changes in flexibility index quantify differences within backbone flexibility, whereas changes in the cooperativity correlation quantify differences within pairwise mechanical couplings. Regardless of metric, the same general conclusions are drawn. That is, small structural perturbations introduced by single point mutations have a frequent and pronounced affect on lysozyme flexibility that can extend over long distances. Specifically, an appreciable change occurs in backbone flexibility for 48% of the residues, and a change in cooperativity occurs in 42% of residue pairs. The average distance from mutation to a site with a change in flexibility is 17–20 Å. Interestingly, the frequency and scale of the changes within single point mutant structures are generally larger than those observed in the hen egg white lysozyme (HEWL) ortholog, which shares 61% sequence identity with human lysozyme. For example, point mutations often lead to substantial flexibility increases within the β-subdomain, which is consistent with experimental results indicating that it is the nucleation site for amyloid formation. However, β-subdomain flexibility within the human and HEWL orthologs is more similar despite the lowered sequence identity. These results suggest compensating mutations in HEWL reestablish desired properties

7-Deazaguanine modifications protect phage DNA from host restriction systems

Genome modifications are central components of the continuous arms race between viruses and their hosts. The archaeosine base (G+), which was thought to be found only in archaeal tRNAs, was recently detected in genomic DNA of Enterobacteria phage 9g and was proposed to protect phage DNA from a wide variety of restriction enzymes. In this study, we identify three additional 2′-deoxy-7-deazaguanine modifications, which are all intermediates of the same pathway, in viruses: 2′-deoxy-7-amido-7-deazaguanine (dADG), 2′-deoxy-7-cyano-7-deazaguanine (dPreQ0) and 2′-deoxy-7- aminomethyl-7-deazaguanine (dPreQ1). We identify 180 phages or archaeal viruses that encode at least one of the enzymes of this pathway with an overrepresentation (60%) of viruses potentially infecting pathogenic microbial hosts. Genetic studies with the Escherichia phage CAjan show that DpdA is essential to insert the 7-deazaguanine base in phage genomic DNA and that 2′-deoxy-7-deazaguanine modifications protect phage DNA from host restriction enzymes

An Essential Difference between the Flavonoids MonoHER and Quercetin in Their Interplay with the Endogenous Antioxidant Network

Antioxidants can scavenge highly reactive radicals. As a result the antioxidants are converted into oxidation products that might cause damage to vital cellular components. To prevent this damage, the human body possesses an intricate network of antioxidants that pass over the reactivity from one antioxidant to another in a controlled way. The aim of the present study was to investigate how the semi-synthetic flavonoid 7-mono-O-(β-hydroxyethyl)-rutoside (monoHER), a potential protective agent against doxorubicin-induced cardiotoxicity, fits into this antioxidant network. This position was compared with that of the well-known flavonoid quercetin. The present study shows that the oxidation products of both monoHER and quercetin are reactive towards thiol groups of both GSH and proteins. However, in human blood plasma, oxidized quercetin easily reacts with protein thiols, whereas oxidized monoHER does not react with plasma protein thiols. Our results indicate that this can be explained by the presence of ascorbate in plasma; ascorbate is able to reduce oxidized monoHER to the parent compound monoHER before oxidized monoHER can react with thiols. This is a major difference with oxidized quercetin that preferentially reacts with thiols rather than ascorbate. The difference in selectivity between monoHER and quercetin originates from an intrinsic difference in the chemical nature of their oxidation products, which was corroborated by molecular quantum chemical calculations. These findings point towards an essential difference between structurally closely related flavonoids in their interplay with the endogenous antioxidant network. The advantage of monoHER is that it can safely channel the reactivity of radicals into the antioxidant network where the reactivity is completely neutralized

Global, regional, and national comparative risk assessment of 79 behavioural, environmental and occupational, and metabolic risks or clusters of risks, 1990-2015: a systematic analysis for the Global Burden of Disease Study 2015

SummaryBackground The Global Burden of Diseases, Injuries, and Risk Factors Study 2015 provides an up-to-date synthesis of the evidence for risk factor exposure and the attributable burden of disease. By providing national and subnational assessments spanning the past 25 years, this study can inform debates on the importance of addressing risks in context. Methods We used the comparative risk assessment framework developed for previous iterations of the Global Burden of Disease Study to estimate attributable deaths, disability-adjusted life-years (DALYs), and trends in exposure by age group, sex, year, and geography for 79 behavioural, environmental and occupational, and metabolic risks or clusters of risks from 1990 to 2015. This study included 388 risk-outcome pairs that met World Cancer Research Fund-defined criteria for convincing or probable evidence. We extracted relative risk and exposure estimates from randomised controlled trials, cohorts, pooled cohorts, household surveys, census data, satellite data, and other sources. We used statistical models to pool data, adjust for bias, and incorporate covariates. We developed a metric that allows comparisons of exposure across risk factors—the summary exposure value. Using the counterfactual scenario of theoretical minimum risk level, we estimated the portion of deaths and DALYs that could be attributed to a given risk. We decomposed trends in attributable burden into contributions from population growth, population age structure, risk exposure, and risk-deleted cause-specific DALY rates. We characterised risk exposure in relation to a Socio-demographic Index (SDI). Findings Between 1990 and 2015, global exposure to unsafe sanitation, household air pollution, childhood underweight, childhood stunting, and smoking each decreased by more than 25%. Global exposure for several occupational risks, high body-mass index (BMI), and drug use increased by more than 25% over the same period. All risks jointly evaluated in 2015 accounted for 57·8% (95% CI 56·6–58·8) of global deaths and 41·2% (39·8–42·8) of DALYs. In 2015, the ten largest contributors to global DALYs among Level 3 risks were high systolic blood pressure (211·8 million [192·7 million to 231·1 million] global DALYs), smoking (148·6 million [134·2 million to 163·1 million]), high fasting plasma glucose (143·1 million [125·1 million to 163·5 million]), high BMI (120·1 million [83·8 million to 158·4 million]), childhood undernutrition (113·3 million [103·9 million to 123·4 million]), ambient particulate matter (103·1 million [90·8 million to 115·1 million]), high total cholesterol (88·7 million [74·6 million to 105·7 million]), household air pollution (85·6 million [66·7 million to 106·1 million]), alcohol use (85·0 million [77·2 million to 93·0 million]), and diets high in sodium (83·0 million [49·3 million to 127·5 million]). From 1990 to 2015, attributable DALYs declined for micronutrient deficiencies, childhood undernutrition, unsafe sanitation and water, and household air pollution; reductions in risk-deleted DALY rates rather than reductions in exposure drove these declines. Rising exposure contributed to notable increases in attributable DALYs from high BMI, high fasting plasma glucose, occupational carcinogens, and drug use. Environmental risks and childhood undernutrition declined steadily with SDI; low physical activity, high BMI, and high fasting plasma glucose increased with SDI. In 119 countries, metabolic risks, such as high BMI and fasting plasma glucose, contributed the most attributable DALYs in 2015. Regionally, smoking still ranked among the leading five risk factors for attributable DALYs in 109 countries; childhood underweight and unsafe sex remained primary drivers of early death and disability in much of sub-Saharan Africa. Interpretation Declines in some key environmental risks have contributed to declines in critical infectious diseases. Some risks appear to be invariant to SDI. Increasing risks, including high BMI, high fasting plasma glucose, drug use, and some occupational exposures, contribute to rising burden from some conditions, but also provide opportunities for intervention. Some highly preventable risks, such as smoking, remain major causes of attributable DALYs, even as exposure is declining. Public policy makers need to pay attention to the risks that are increasingly major contributors to global burden. Funding Bill & Melinda Gates Foundation

Identification of host proteins interacting with Toxoplasma gondii GRA15 (TgGRA15) by yeast two-hybrid system

Background Toxoplasma gondii, an obligate intracellular protozoan parasite, possesses the remarkable ability to co-opt host cell machinery in order to maintain its intracellular survival. This parasite can modulate signaling pathways of its host through the secretion of polymorphic effector proteins localized in the rhoptry and dense granule organelles. One of such effectors is T. gondii type II-specific dense granule protein 15, TgGRA15, which activates NF-κB pathway. The aim of the present study was to identify the host interaction partner proteins of TgGRA15. Methods We screened a yeast two-hybrid mouse cDNA library using TgGRA15 as the bait. TgGRA15 (PRU strain, Type II) was cloned into the pGBKT7 vector and expressed in the Y2HGold yeast strain. Then, the bait protein expression was validated by western blotting analysis, followed by auto-activation and toxicity tests in comparison with control (Y2HGold yeast strain transformed with empty pGBKT7 vector). Results This screening led to the identification of mouse Luzp1 and AW209491 as host binding proteins that interact with TgGRA15. Luzp1 contains three nuclear localizing signals and is involved in regulating a subset of host non-coding RNA genes. Conclusions These findings reveal, for the first time, new host cell proteins interacting with TgGRA15. The identification of these cellular targets and the understanding of their contribution to the host-pathogen interaction may serve as the foundation for novel therapeutic and prevention strategies against T. gondii infection

RNAi-mediated silencing of the Bmi-1 gene causes growth inhibition and enhances doxorubicin-induced apoptosis in MCF-7 cells

The oncogene Bmi-1 is a member of the Polycomb group gene family. Its expression is found to be greatly increased in a number of malignant tumors including breast cancer. This could suggest Bmi-1 as a potent therapeutic target. In this study, RNAi was introduced to down-regulate the expression of Bmi-1 in a highly malignant breast adenocarcinoma cell line, MCF-7. A thorough study of the biological behavior and chemosensitivity changes of the MCF-7 cells was carried out in context to the therapeutic potential of Bmi-1. The results obtained indicated that siRNA targeting of Bmi-1 could lead to an efficient and specific inhibition of endogenous Bmi-1 activity. The mRNA and protein expression of Bmi-1 were determined by RT-PCR and Western blot, respectively. Furthermore, silencing of Bmi-1 resulted in a drastic inhibition of the growth of MCF-7 cells as well as G1 /S phase transition. The number of target cells was found to increase in phase G 0 /G 1 and decrease in the S phase, but no increase in the basal level of apoptosis was noticed. On the other hand, a reduction in the expression of cyclin D1 and an increase in the expression of p21 were also noticed. Silencing of Bmi-1 made the MCF-7 cells more sensitive to the chemotherapeutic agent doxorubicin and induced a significantly higher percentage of apoptotic cells. Here, we report on a study regarding the RNAi-mediated silencing of the Bmi-1 gene in breast cancer