Analyzing the Catalytic Role of Asp97 in the Methionine Aminopeptidase from \u3cem\u3eEscherichia coli\u3c/em\u3e

An active site aspartate residue, Asp97, in the methionine aminopeptidase (MetAPs) from Escherichia coli (EcMetAP-I) was mutated to alanine, glutamate, and asparagine. Asp97 is the lone carboxylate residue bound to the crystallographically determined second metal-binding site in EcMetAP-I. These mutant EcMetAP-I enzymes have been kinetically and spectroscopically characterized. Inductively coupled plasma–atomic emission spectroscopy analysis revealed that 1.0 ± 0.1 equivalents of cobalt were associated with each of the Asp97-mutated EcMetAP-Is. The effect on activity after altering Asp97 to alanine, glutamate or asparagine is, in general, due to a ∼ 9000-fold decrease in kca towards Met-Gly-Met-Met as compared to the wild-type enzyme. The Co(II) d–d spectra for wild-type, D97E and D97A EcMetAP-I exhibited very little difference in form, in each case, between the monocobalt(II) and dicobalt(II) EcMetAP-I, and only a doubling of intensity was observed upon addition of a second Co(II) ion. In contrast, the electronic absorption spectra of [Co_(D97N EcMetAP-I)] and [CoCo(D97N EcMetAP-I)] were distinct, as were the EPR spectra. On the basis of the observed molar absorptivities, the Co(II) ions binding to the D97E, D97A and D97N EcMetAP-I active sites are pentacoordinate. Combination of these data suggests that mutating the only nonbridging ligand in the second divalent metal-binding site in MetAPs to an alanine, which effectively removes the ability of the enzyme to form a dinuclear site, provides a MetAP enzyme that retains catalytic activity, albeit at extremely low levels. Although mononuclear MetAPs are active, the physiologically relevant form of the enzyme is probably dinuclear, given that the majority of the data reported to date are consistent with weak cooperative binding

Jeremiah's kings : a study of the book's treatment of the monarchy, with special reference to chapters 21-24

Starting from an analysis of approaches to the book of Jeremiah adopted towards the end of the last century, this thesis enquires what I ight is thrown on its redactional history by the way in which the kings purportedly reigning during the prophet's ministry, and also David and Nebuchadnezzar, are treated in the book. One objective is to see where the book should be placed in the spectrum of conclusions arrived at in recent years, supposing that the commentary of W. L.Holladay stands at one end of this spectrum with his belief in the historical reliability of most of the information contained in the book, and that ofR.P.Carroll at the other with his scepticism from this point of view. The starting point for this enquiry is the collection of material about kings in Jeremiah 21-2t, but succeeding chapters of the thesis, dealing in turn with those concerned, namely losiah, lehoahaz, lehoiakim, lehoiachin and Zedekiah, examine also other parts of the book where they are mentioned. David and Nebuchadnezzar are both seen to be important figures in Jeremiah with regard to the question of the monarchy, and both very relevant to the argument. A distinctive stance is taken with regard to the obscure figure of Zerubbabel. The main conclusion is that throughout the book of Jeremiah there is evidence of a lengthy history of redaction, not only in the case of alterations made by scribes for no significant reason, but also in many passages where changes have been made from contrasting religio-political points of view, not least with regard to the understanding of the monarchy itself A final chapter offers reflections on the question how, taking serious account of its complex and turbulent redactional history, the book of Jeremiah may be read today as Christian scripture

Understanding the mechanisms of IGF2 gene regulation in hepatocellular carcinoma cells

Hepatocellular carcinoma (HCC) is a leading cause of cancer-related deaths worldwide. HCC has a very well studied etiology, and is associated with chronic hepatic viral infections (hepatitis viruses B and C), alcohol abuse, or other causes of chronic liver damage. Currently, tumor resection and liver transplantation are the only potentially curative treatments available for HCC. However, the presence of extra-hepatic invasion and metastasis makes patients ineligible for these treatments. High IGF2 levels are associated with metastatic HCC, and we recently showed that IGF2-induced signaling through Igf1R stimulates the invasiveness and metastatic phenotype of HCC cells. However, the precise mechanisms by which IGF2 expression is enhanced in HCC are not well understood. IGF2 is an imprinted gene normally expressed from the paternal allele. Loss of imprinting, which activates the normally silent maternal allele, has been implicated as an epigenetic marker for the enhanced risk of human cancer. However, many HCCs that display elevated IGF2 expression levels retain a normal imprinting pattern. Therefore, additional gene regulation mechanisms must also influence IGF2 expression in HCC. Hypothesis: Long-range genomic interactions are important for the regulation of IGF2 gene expression, and alterations in these long-range interactions lead to elevated IGF2 gene expression in HCC. To address this hypothesis I have utilized chromosome conformation capture carbon copy (5C) technology to elucidate long-range interactions involving the IGF2 promoters in a normal hepatocyte cell line, THLE-2, and an HCC cell line HepG2

Rodenticide Exposure Among Endangered Kit Foxes Relative to Habitat Use in an Urban Landscape

Endangered San Joaquin kit foxes (Vulpes macrotis mutica) inhabiting Bakersfield, California exhibit a high incidence of exposure to anticoagulant rodenticides (ARs). We examined kit fox habitat use in an effort to determine potential sources of AR exposure. Kit fox capture, den, night, and mortality locations were assigned to one of 10 habitat categories. Using all available locations, foxes that tested positive for second generation anticoagulant rodenticides (SGARs) were located more frequently on golf courses while those testing negative were located more frequently in commercial areas. Foxes that tested positive for first generation anticoagulant rodenticides (FGARs) were located more frequently in industrial areas while those testing negative were located more frequently on golf courses. Based on night locations (when foxes are foraging), foxes that tested positive for SGARs were found more frequently in undeveloped and golf course habitats. Foxes that tested positive for FGARs were found more frequently in undeveloped, campus, and industrial habitats. Although available data were not sufficient to identify specific point-sources of AR exposure for foxes, golf courses appeared to be used more frequently by foxes exposed to SGARs. However, sources of exposure likely are abundant and widespread in the urban environment. Based on the results of this study, we recommend (1) investigating patterns of AR use in Bakersfield, (2) conducting an outreach program to emphasize the risk from ARs to kit foxes and other wildlife, and (3) continuing to monitor the incidence and patterns of AR exposure among kit foxes in Bakersfield

Intergroup emotional exchange: Ingroup guilt and outgroup anger increase resource allocation in trust games

Intergroup exchanges are an integral part of social life but are compromised when one group pursues its interests at another group’s expense. The present research investigates whether expressing emotion can mitigate the negative consequences of such actions. We examine how emotions communicated by either an ingroup or outgroup member following an ingroup member’s breach of trust affect other ingroup members’ feelings of guilt and pride, and subsequent allocation of resources. In both studies, groups of participants played a two-round trust game with another group. In round one, they observed a member of their own group failing to reciprocate a trusting move by the outgroup. In Study 1 (N = 85), an outgroup member then communicated anger or disappointment, whereas in Study 2 (N = 164), an ingroup member then communicated happiness or guilt. Comparisons with no-emotion control conditions revealed that expressions of outgroup anger and ingroup guilt increased participants’ allocations to an outgroup member in round two. The effect of an outgroup member’s anger expression was mediated by participants’ diminished feelings of pride about the ingroup action, whereas the effect of an ingroup member’s guilt expression was mediated by participants’ own feelings of guilt. Taken together, these findings support a social appraisal approach and highlight the roles that pride and guilt can play in shaping intergroup resource allocations

Severe anemia in Malawian children

Background Severe anemia is a major cause of sickness and death in African children, yet the causes of anemia in this population have been inadequately studied. Methods We conducted a case-control study of 381 preschool children with severe anemia (hemoglobin concentration, <5.0 g per deciliter) and 757 preschool children without severe anemia in urban and rural settings in Malawi. Causal factors previously associated with severe anemia were studied. The data were examined by multivariate analysis and structural equation modeling. Results Bacteremia (adjusted odds ratio, 5.3; 95% confidence interval [CI], 2.6 to 10.9), malaria (adjusted odds ratio, 2.3; 95% CI, 1.6 to 3.3), hookworm (adjusted odds ratio, 4.8; 95% CI, 2.0 to 11.8), human immunodeficiency virus infection (adjusted odds ratio, 2.0; 95% CI, 1.0 to 3.8), the G6PD(sup -202/-376) genetic disorder (adjusted odds ratio, 2.4; 95% CI, 1.3 to 4.4), vitamin A deficiency (adjusted odds ratio, 2.8; 95% CI, 1.3 to 5.8), and vitamin B(sub 12) deficiency (adjusted odds ratio, 2.2; 95% CI, 1.4 to 3.6) were associated with severe anemia. Folate deficiency, sickle cell disease, and laboratory signs of an abnormal inflammatory response were uncommon. Iron deficiency was not prevalent in case patients (adjusted odds ratio, 0.37; 95% CI, 0.22 to 0.60) and was negatively associated with bacteremia. Malaria was associated with severe anemia in the urban site (with seasonal transmission) but not in the rural site (where malaria was holoendemic). Seventy-six percent of hookworm infections were found in children under 2 years of age. Conclusions There are multiple causes of severe anemia in Malawian preschool children, but folate and iron deficiencies are not prominent among them. Even in the presence of malaria parasites, additional or alternative causes of severe anemia should be considere

Research Article (New England Journal of Medicine) Severe anemia in Malawian children

Background: Severe anemia is a major cause of sickness and death in African children, yet the causes of anemia in this population have been inadequately studied.Methods: We conducted a case–control study of 381 preschool children with severe anemia (hemoglobin concentration, &lt;5.0 g per deciliter) and 757 preschool children without severe anemia in urban and rural settings in Malawi. Causal factors  previously associated with severe anemia were studied. The data were examined by multivariate analysis and structural equation modeling.Results: Bacteremia (adjusted odds ratio, 5.3; 95% confidence interval [CI], 2.6 to 10.9), malaria (adjusted odds ratio, 2.3; 95% CI, 1.6 to 3.3), hookworm (adjusted odds ratio, 4.8; 95% CI, 2.0 to 11.8), human immunodeficiency virus infection (adjusted odds ratio, 2.0; 95% CI, 1.0 to 3.8), the G6PD−202/−376 genetic disorder (adjusted odds ratio, 2.4; 95% CI, 1.3 to 4.4), vitamin A deficiency (adjusted odds ratio, 2.8; 95% CI, 1.3 to 5.8), and vitamin B12 deficiency (adjusted odds ratio, 2.2; 95% CI, 1.4 to 3.6) were associated with severe anemia. Folate deficiency, sickle cell disease, and laboratory signs of an abnormal  inflammatory response were uncommon. Iron deficiency was not prevalent in case patients (adjusted odds ratio, 0.37; 95% CI, 0.22 to 0.60) and was negatively associated with bacteremia. Malaria was associated with severe anemia in the urban site (with seasonal transmission) but not in the rural site (where malaria was holoendemic). Seventy-six percent of hookworm infections were found in children under 2 years of age.Conclusions: There are multiple causes of severe anemia in Malawian preschool children, but folate and iron deficiencies are not prominent among them. Even in the presence of malaria parasites, additional or alternative causes of severe anemia should be considered

Use of digital measurement of medication adherence and lung function to guide the management of uncontrolled asthma (INCA Sun):a multicentre, single-blinded, randomised clinical trial

BACKGROUND: The clinical value of using digital tools to assess adherence and lung function in uncontrolled asthma is not known. We aimed to compare treatment decisions guided by digitally acquired data on adherence, inhaler technique, and peak flow with existing methods.METHODS: A 32-week prospective, multicentre, single-blinded, parallel, randomly controlled trial was done in ten severe asthma clinics across Ireland, Northern Ireland, and England. Participants were 18 years or older, had uncontrolled asthma, asthma control test (ACT) score of 19 or less, despite treatment with high-dose inhaled corticosteroids, and had at least one severe exacerbation in the past year despite high-dose inhaled corticosteroids. Patients were randomly assigned in a 1:1 ratio to the active group or the control group, by means of a computer-generated randomisation sequence of permuted blocks of varying sizes (2, 4, and 6) stratified by fractional exhaled nitric oxide (FeNO) concentration and recruitment site. In the control group, participants were masked to their adherence and errors in inhaler technique data. A statistician masked to study allocation did the statistical analysis. After a 1-week run-in period, both groups attended three nurse-led education visits over 8 weeks (day 7, week 4, and week 8) and three physician-led treatment adjustment visits at weeks 8, 20, and 32. In the active group, treatment adjustments during the physician visits were informed by digital data on inhaler adherence, twice daily digital peak expiratory flow (ePEF), patient-reported asthma control, and exacerbation history. Treatment was adjusted in the control group on the basis of pharmacy refill rates (a measure of adherence), asthma control by ACT questionnaire, and history of exacerbations and visual management of inhaler technique. Both groups used a digitally enabled Inhaler Compliance Assessment (INCA) and PEF. The primary outcomes were asthma medication burden measured as proportion of patients who required a net increase in treatment at the end of 32 weeks and adherence rate measured in the last 12 weeks by area under the curve in the intention-to-treat population. The safety analyses included all patients who consented for the trial. The trial is registered with ClinicalTrials.gov, NCT02307669 and is complete.FINDINGS: Between Oct 25, 2015, and Jan 26, 2020, of 425 patients assessed for eligibility, 220 consented to participate in the study, 213 were randomly assigned (n=108 in the active group; n=105 in the control group) and 200 completed the study (n=102 in the active group; n=98 in the control group). In the intention-to-treat analysis at week 32, 14 (14%) active and 31 (32%) control patients had a net increase in treatment compared with baseline (odds ratio [OR] 0·31 [95% CI 0·15-0·64], p=0·0015) and 11 (11%) active and 21 (21%) controls required add-on biological therapy (0·42 [0·19-0·95], p=0·038) adjusted for study site, age, sex, and baseline FeNO. Three (16%) of 19 active and 11 (44%) of 25 control patients increased their medication from fluticasone propionate 500 μg daily to 1000 μg daily (500 μg twice a day; adjusted OR 0·23 [0·06-0·87], p=0·026). 26 (31%) of 83 active and 13 (18%) of 73 controls reduced their medication from fluticasone propionate 1000 μg once daily to 500 μg once daily (adjusted OR 2·43 [1·13-5·20], p=0·022. Week 20-32 actual mean adherence was 64·9% (SD 23·5) in the active group and 55·5% (26·8) in the control group (between-group difference 11·1% [95% CI 4·4-17·9], p=0·0012). A total of 29 serious adverse events were recorded (16 [55%] in the active group, and 13 [45%] in the control group), 11 of which were confirmed as respiratory. None of the adverse events reported were causally linked to the study intervention, to the use of salmeterol-fluticasone inhalers, or the use of the digital PEF or INCA.INTERPRETATION: Evidence-based care informed by digital data led to a modest improvement in medication adherence and a significantly lower treatment burden.FUNDING: Health Research Board of Ireland, Medical Research Council, INTEREG Europe, and an investigator-initiated project grant from GlaxoSmithKline.</p