Cloning and expression of a thermostable α-galactosidase from the thermophilic fungus Talaromyces emersonii in the methylotrophic yeast Pichia pastoris

The first gene (alpha-gal1) encoding an extracellular alpha-Dgalactosidase from the thermophilic fungus Talaromyces emersonii was cloned and characterized. The alpha-gal1 gene consisted of an open reading frame of 1,792 base pairs interrupted by six introns that encoded a mature protein of 452 amino acids, including a 24 amino acid secretory signal sequence. The translated protein had highest identity with other fungal alpha-galactosidases belonging to glycosyl hydrolase family 27. The alpha-gal1 gene was overexpressed as a secretory protein with an N-terminal histidine tag in the methylotrophic yeast Pichia pastoris. Recombinant alpha-Gal1 was secreted into the culture medium as a monomeric glycoprotein with a maximal yield of 10.75 mg/l and purified to homogeneity using Hisbinding nickel-agarose affinity chromatography. The purified enzyme was maximally active at 70 degrees C, pH 4.5, and lost no activity over 10 days at 50 degrees C. alpha-Gal1 followed Michaelis-Menten kinetics (Vmax of 240.3 micronM/min/mg, Km of 0.294 mM) and was inhibited competitively by galactose (Km obs of 0.57 mM, Ki of 2.77 mM). The recombinant T. emersonii alpha-galactosidase displayed broad substrate preference, being active on both oligo- and polymeric substrates, yet had strict specificity for the alpha-galactosidic linkage. Owing to its substrate preference and noteworthy stability, alpha-Gal1 is of particular interest for possible biotechnological applications involving the processing of plant materials.J.S. thanks her supervisor at the University of Jyvaskyla, Emily Knott, for permission to perform her Masters degree research while on exchange at NUT Galway, and for constructive comments on the manuscript. J.S. and A.G. both received scholarships under EU Erasmus/Socrates bilateral agreement. Funding for this research was provided in part to M.G.T. under the National Development Plan, through the Food Institutional Research Measure, administered by the Department of Agriculture, Fisheries and Food, Ireland

Physiological and metabolic characteristics of elite tug of war athletes

Objective—To determine the aerobic power ([Image: see text] O(2)MAX), body composition, strength, muscular power, flexibility, and biochemical profile of an elite international squad of tug of war athletes. Methods—Sixteen male competitors (mean (SEM) age 34 (2) years) were evaluated in a laboratory. For comparative purposes, data were analysed relative to normative data for our centre and to a group of 20 rugby forwards from the Irish international squad. Results—The tug of war participants were lighter (83.6 (3.0) v 104.4 (1.8) kg, p<0.0001) and had less lean body mass (69.4 (2.1) v 86.2 (1.2) kg) than the rugby players and had lower than normal body fat (16.7 (0.9)%); all values are mean (SEM). Aerobic power measured during a treadmill test was 55.8 (1.6) ml/kg/min for the tug of war participants compared with 51.1 (1.4) ml/kg/min for the rugby forwards (p<0.03). A composite measure of strength derived from (sum of dominant and non-dominant grip strength and back strength)/lean body mass yielded a strength/mass ratio that was 32% greater (p<0.0001) for the tug of war group than the rugby group. Dynamic leg power was lower for the tug of war group than the rugby forwards (4659.8 (151.6) v 6198.2 (105) W respectively; p<0.0001). Leg flexibility was 25.4 (2.0) cm for the tug of war group. Back flexibility was 28.6 (1.4) cm which was lower (p<0.02) than the rugby forwards 34.2 (1.5) cm. Whereas blood chemistry and haematology were normal, packed cell volume, haemoglobin concentration, and erythrocyte volume were lower in the tug of war group than in the rugby players (p<0.05). All three haematological measures correlated with muscle mass (packed cell volume, r(2) = 0.37, p<0.0001; haemoglobin concentration, r(2) = 0.13, p<0.05; erythrocyte volume, r(2) = 0.21, p<0.01). Conclusions—The data indicate that international level tug of war participants have excellent strength and above average endurance relative to body size, but have relatively low explosive leg power and back flexibility. The data provide reference standards for the sport and may be useful for monitoring and evaluating current and future participants. Key Words: tug of war; body composition; [Image: see text] O(2)MAX; strength; power; flexibilit

Book reviews

Teaching/Communication/Extension/Profession,

Primary and Secondary Control as Antecedents to the Dark Traits in Predicting Attraction to Hacking Behavior

The current study examines the relationship between the need for control, the Dark Triad personality traits, and hacking intent. We surveyed 523 individuals using a scenario design and investigated the role of both primary and secondary control as antecedents to Machiavellianism, psychopathy, and narcissism leading to both white- (i.e., ethical) and black-hat hacking interest. Our findings suggest that primary control is a significant antecedent to all three dark personality traits such that a higher intrinsic need for control is positively associated with Machiavellianism, narcissism, and psychopathy. Secondary control, however, has comparatively different effects on dark personality traits, demonstrating a negative effect only on psychopathy. Both Machiavellianism and psychopathy predicted both white- and black-hat hacking interests along with the perceived probability of apprehension. Overall, our findings suggest that primary control drives all three dark personality traits, yet only two of the three dark personality traits – Machiavellianism and psychopathy – are related to hacking interest

Update on the Surgical Trial in Lobar Intracerebral Haemorrhage (STICH II):Statistical analysis plan

<p>Abstract</p> <p>Background</p> <p>Previous studies had suggested that the outcome for patients with spontaneous lobar intracerebral haemorrhage (ICH) and no intraventricular haemorrhage (IVH) might be improved with early evacuation of the haematoma. The Surgical Trial in Lobar Intracerebral Haemorrhage (STICH II) set out to establish whether a policy of earlier surgical evacuation of the haematoma in selected patients with spontaneous lobar ICH would improve outcome compared to a policy of initial conservative treatment. It is an international, multi-centre, prospective randomised parallel group trial of early surgery in patients with spontaneous lobar ICH. Outcome is measured at six months via a postal questionnaire.</p> <p>Results</p> <p>Recruitment to the study began on 27 November 2006 and closed on 15 August 2012 by which time 601 patients had been recruited. The protocol was published in <it>Trials</it> (<url>http://www.trialsjournal.com/content/12/1/124/</url>). This update presents the analysis plan for the study without reference to the unblinded data. The trial data will not be unblinded until after follow-up is completed in early 2013. The main trial results will be presented in spring 2013 with the aim to publish in a peer-reviewed journal at the same time.</p> <p>Conclusion</p> <p>The data from the trial will provide evidence on the benefits and risks of early surgery in patients with lobar ICH.</p> <p>Trial registration</p> <p>ISRCTN: ISRCTN22153967</p

Further development and flight test of an autonomous precision landing system using a parafoil

NASA Dryden Flight Research Center and NASA Johnson Space Center are jointly conducting a phased program to determine the feasibility of the autonomous recovery of a spacecraft using a ram-air parafoil system for the final stages of entry from space to a precision landing. The feasibility is being studied using a flight model of a spacecraft in the generic shape of a flattened biconic that weighs approximately 120 lb and is flown under a commercially available ram-air parafoil. Key components of the vehicle include the global positioning system (GPS) guidance for navigation, a flight control computer, an electronic compass, a yaw rate gyro, and an onboard data recorder. A flight test program is being used to develop and refine the vehicle. The primary flight goal is to demonstrate autonomous flight from an altitude of 3,000 m (10,000 ft) with a lateral offset of 1.6 km (1.0 mi) to a precision soft landing. This paper summarizes the progress to date. Much of the navigation system has been tested, including a heading tracker that was developed using parameter estimation techniques and a complementary filter. The autoland portion of the autopilot is still in development. The feasibility of conducting the flare maneuver without servoactuators was investigated as a means of significantly reducing the servoactuator rate and load requirements

Collaborative College Leadership

With the collaborative leadership style of a new dean, the Dean’s Executive Council (associate deans, school directors, chair) in the BGSU College of Education and Human Development transformed operations of the team and our work with college faculty. This has reenergized the college and garnered substantial faculty support and engagement

6′,7′‐Dihydroxybergamottin in grapefruit juice and Seville orange juice: Effects on cyclosporine disposition, enterocyte CYP3A4, and P‐glycoprotein

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/109869/1/cptclpt1999363.pd

A Novel Fluorescent Clinical Method to Rapidly Quantify Plasma Volume

Objectives To determine the performance of a rapid fluorescent indicator technique for measuring plasma volume (PV). Methods This was an open-label, observational evaluation of a two-component intravenous visible fluorescent dye technique to rapidly measure PV in 16 healthy subjects and 16 subjects with chronic kidney disease (8 stage 3 and 8 stage 4 CKD), at 2 clinical research sites. The method consisted of a single intravenous injection of 12 mg of a large 150-kDa carboxy-methyl dextran conjugated to a fluorescent rhodamine-derived dye as the PV marker (PVM), and 35 mg of a small 5-kDa carboxy-methyl dextran conjugated to fluorescein, the renal clearance marker. Dye concentrations were quantified 15 min after the injections for initial PV measurements using the indicator-dilution principle. Additional samples were taken over 8 h to evaluate the stability of the PVM as a determinant of PV. Blood volumes (BV) were calculated based on PV and the subject’s hematocrit. Pharmacokinetic parameters were calculated from the plasma concentration data taken over several days using noncompartmental methods (Phoenix WinNonlin®). Linear correlation and Bland-Altman plots were used to compare visible fluorescent injectate-measured PV compared to Nadler’s formula for estimating PV. Finally, 8 healthy subjects received 350 mL infusion of a 5% albumin solution in normal saline over 30 min and a repeat PV determination was then carried out. Results PV and BV varied according to weight and body surface area, with PV ranging from 2,115 to 6,234 mL and 28.6 to 41.9 mL/kg when weight adjusted. Both parameters were stable for > 6 h with repeated plasma measurements of the PVM. There was no difference between healthy subjects and CKD subjects. Overall, there was general agreement with Nadler’s estimation formula for the mean PV in subjects. A 24-h repeat dose measurement in 8 healthy subjects showed PV variability of 98 ± 121 mL (mean = 3.8%). Additionally, following an intravenous bolus of 350 mL of a 5% albumin solution in normal saline in 8 healthy subjects, the mean (SD) measured increase in PV was 356 (±50.0) mL post-infusion. There were no serious adverse events reported during the study. Conclusions This minimally invasive fluorescent dye approach safely allowed for rapid, accurate, and reproducible determination of PV, BV, and dynamic monitoring of changes following fluid administration