ENZYMES: Catalysis, Kinetics and Mechanisms

Onemarvelsattheintricate designoflivingsystems,andwecannotbutwonderhow life originated on this planet. Whether ?rst biological structures emerged as the selfreproducing genetic templates (genetics-?rst origin of life) or the metabolic universality preceded the genome and eventually integrated it (metabolism-?rst origin of life) is still a matter of hot scienti?c debate. There is growing acceptance that the RNA world came ?rst – as RNA molecules can perform both the functions of information storage and catalysis. Regardless of which view eventually gains acceptance, emergence of catalytic phenomena is at the core of biology. The last century has seen an explosive growth in our understanding of biological systems. The progression has involved successive emphasis on taxonomy ! physiology ! biochemistry ! molecular biology ! genetic engineering and ?nally the large-scale study of genomes. The ?eld of molecular biology became largely synonymous with the study of DNA – the genetic material. Molecular biology however had its beginnings in the understanding of biomolecular structure and function. Appreciationofproteins,catalyticphenomena,andthefunctionofenzymeshadalargeroleto play in the progress of modern biology

Studies on Aspergillus niger glutamine synthetase: regulation of enzyme levels by nitrogen sources and identification of active site residues

The specific activity of glutamine synthetase (L-glutamate: ammonia ligase, EC 6.3.1.2) in surface grown Aspergillus niger was increased 3-5 fold when grown on L-glutamate or potassium nitrate, compared to the activity obtained on ammonium chloride. The levels of glutamine synthetase was regulated by the availability of nitrogen source like NH4+ , and further, the enzyme is repressed by increasing concentrations of NH4+. In contrast to other micro-organisms, the Aspergillus niger enzyme was neither specifically inactivated by NH4+ or L-glutamine nor regulated by covalent modification. Glutamine synthetase from Aspergillus niger was purified to homogenity. The native enzyme is octameric with a molecular weight of 385,000±25,000. The enzyme also catalyses Mn2+ or Mg2+-dependent synthetase and Mn2+-dependent transferase activity. Aspergillus niger glutamine synthetase was completely inactivated by two mol of phenylglyoxal and one mol of N-ethylmaleimide with second order rate constants of 3.8 M-1 min-1 and 760 M-1 min-1 respectively. Ligands like Mg. ATP, Mg. ADP, Mg. AMP, L-glutamate NH4+, Mn2+ protected the enzyme against inactivation. The pattern of inactivation and protection afforded by different ligands against N-ethylamaleimide and phenylglyoxal was remarkably similar. These results suggest that metal ATP complex acts as a substrate and interacts with an arginine ressidue at the active site. Further, the metal ion and the free nucleotide probably interact at other sites on the enzyme affecting the catalytic activity

Isolation of genomic DNA from acetone-dried Aspergillus mycelia

A simple acetone-drying protocol was adopted to replace the lyophilization step while isolating genomic DNA from Aspergillus mycelia. This DNA is suitable for PCR, restriction enzyme digestion and Southern blot analysis with digoxigenin-labeled DNA probes. Acetone drying/ preservation can be a useful method in the molecular analysis of fungal DNA samples

Anomaly Detection for Vision-based Railway Inspection

none7nomixedRiccardo Gasparini; Stefano Pini; Guido Borghi; Giuseppe Scaglione; Simone Calderara; Eugenio Fedeli; Rita CucchiaraRiccardo Gasparini; Stefano Pini; Guido Borghi; Giuseppe Scaglione; Simone Calderara; Eugenio Fedeli; Rita Cucchiar

Prevalence of Allergic Disorders among Primary School-Aged Children in Madinah, Saudi Arabia: Two-Stage Cross-Sectional Survey

There are limited data on the epidemiology of allergic disorders in Saudi Arabia. Such data are needed for, amongst other things, helping to plan service provision at a time when there is considerable investment taking place in national healthcare development. We sought to estimate the prevalence of atopic eczema, allergic rhinitis and asthma in primary school children in Madinah, Saudi Arabia.We conducted a two-stage cross-sectional survey of schoolchildren in Madinah. Children were recruited from 38 randomly selected schools. Questionnaires were sent to the parents of all 6,139 6-8 year old children in these schools. These parental-completed questionnaires incorporated questions from the International Study of Asthma and Allergies in Childhood (ISAAC), which had previously been validated for use in Arab populations. We undertook descriptive analyses, using the Generalized Estimating Equation (GEE) to calculate 95% confidence intervals. The overall response rate was 85.9% (n = 5,188), 84.6% for girls and 86.2% for boys, respectively. Overall, parents reported symptoms suggestive of a history of eczema in 10.3% (95%CI 9.4, 11.4), rhinitis in 24.2% (95%CI 22.3, 26.2) and asthma in 23.6% (95%CI 21.3, 26.0) of children. Overall, 41.7% (95%CI 39.1, 44.4) of children had symptoms suggestive of at least one allergic disorder, with a substantial minority manifesting symptoms indicative of co-morbid allergic disease. Comparison of these symptom-based prevalence estimates with reports of clinician-diagnosed disease suggested that the majority of children with eczema and asthma had been diagnosed, but only a minority (17.4%) of children had been diagnosed with rhinitis. International comparisons indicated that children in Madinah have amongst the highest prevalence of allergic problems in the world.Symptoms indicative of allergic disease are very common in primary school-aged children in Madinah, Saudi Arabia, with figures comparable to the highest risk regions in the world

Macrofossil evidence for a rapid and severe Cretaceous–Paleogene mass extinction in Antarctica

Debate continues about the nature of the Cretaceous–Paleogene (K–Pg) mass extinction event. An abrupt crisis triggered by a bolide impact contrasts with ideas of a more gradual extinction involving flood volcanism or climatic changes. Evidence from high latitudes has also been used to suggest that the severity of the extinction decreased from low latitudes towards the poles. Here we present a record of the K–Pg extinction based on extensive assemblages of marine macrofossils (primarily new data from benthic molluscs) from a highly expanded Cretaceous–Paleogene succession: the López de Bertodano Formation of Seymour Island, Antarctica. We show that the extinction was rapid and severe in Antarctica, with no significant biotic decline during the latest Cretaceous, contrary to previous studies. These data are consistent with a catastrophic driver for the extinction, such as bolide impact, rather than a significant contribution from Deccan Traps volcanism during the late Maastrichtian

Effects of antiplatelet therapy on stroke risk by brain imaging features of intracerebral haemorrhage and cerebral small vessel diseases: subgroup analyses of the RESTART randomised, open-label trial

Background Findings from the RESTART trial suggest that starting antiplatelet therapy might reduce the risk of recurrent symptomatic intracerebral haemorrhage compared with avoiding antiplatelet therapy. Brain imaging features of intracerebral haemorrhage and cerebral small vessel diseases (such as cerebral microbleeds) are associated with greater risks of recurrent intracerebral haemorrhage. We did subgroup analyses of the RESTART trial to explore whether these brain imaging features modify the effects of antiplatelet therapy

Adolescent seasonal allergic rhinitis and the impact of health-care professional training:cluster randomised controlled trial of a complex intervention in primary care

BACKGROUND: Seasonal allergic rhinitis is typically poorly managed, particularly in adolescents, in whom it is responsible for considerable morbidity. Our previous work has demonstrated that if poorly controlled this can impair educational performance. AIM: The primary aim of this trial was to assess the impact of a primary care-based professional training intervention on clinical outcomes in adolescents with seasonal allergic rhinitis. METHODS: Cluster trial in which UK general practice staff were randomised to a short, intensive workshop on the evidence-based management of seasonal allergic rhinitis. The primary outcome measure was the change in the validated Rhinoconjunctivitis Quality of Life Questionnaire with Standardized Activities (RQLQ(S)) score between baseline and 6 weeks post intervention (minimal clinically important difference=0.5). Secondary outcome measures of interest included health-care professionals' knowledge and confidence in managing seasonal allergic rhinitis, number of seasonal allergic rhinitis-related consultations, relevant treatments prescribed and symptom scores. RESULTS: Thirty-eight general practices were randomised (20 in the intervention arm) and 246 patients (50.2% males, mean age 15 years) were included in the primary outcome analysis. Health-care professionals' knowledge and confidence of the clinical management of seasonal allergic rhinitis improved. This did not, however, result in clinically or statistically significant improvements in RQLQ(S): -0.15, (95% confidence interval, -0.5 to +0.2). There were no differences in consultation frequency, treatments issued for seasonal allergic rhinitis or symptom scores. CONCLUSIONS: Although associated with increases in professionals' self-assessed confidence and understanding of seasonal allergic rhinitis management, this intensive training workshop did not translate into improvements in adolescents' disease-specific quality of life or a reduction in rhinitis symptoms

Status Update and Interim Results from the Asymptomatic Carotid Surgery Trial-2 (ACST-2)

Objectives: ACST-2 is currently the largest trial ever conducted to compare carotid artery stenting (CAS) with carotid endarterectomy (CEA) in patients with severe asymptomatic carotid stenosis requiring revascularization. Methods: Patients are entered into ACST-2 when revascularization is felt to be clearly indicated, when CEA and CAS are both possible, but where there is substantial uncertainty as to which is most appropriate. Trial surgeons and interventionalists are expected to use their usual techniques and CE-approved devices. We report baseline characteristics and blinded combined interim results for 30-day mortality and major morbidity for 986 patients in the ongoing trial up to September 2012. Results: A total of 986 patients (687 men, 299 women), mean age 68.7 years (SD ± 8.1) were randomized equally to CEA or CAS. Most (96%) had ipsilateral stenosis of 70-99% (median 80%) with contralateral stenoses of 50-99% in 30% and contralateral occlusion in 8%. Patients were on appropriate medical treatment. For 691 patients undergoing intervention with at least 1-month follow-up and Rankin scoring at 6 months for any stroke, the overall serious cardiovascular event rate of periprocedural (within 30 days) disabling stroke, fatal myocardial infarction, and death at 30 days was 1.0%. Conclusions: Early ACST-2 results suggest contemporary carotid intervention for asymptomatic stenosis has a low risk of serious morbidity and mortality, on par with other recent trials. The trial continues to recruit, to monitor periprocedural events and all types of stroke, aiming to randomize up to 5,000 patients to determine any differential outcomes between interventions. Clinical trial: ISRCTN21144362. © 2013 European Society for Vascular Surgery. Published by Elsevier Ltd. All rights reserved