Recombinant production of the therapeutic peptide lunasin

<p>Abstract</p> <p>Background</p> <p>Lunasin is a chemopreventive peptide produced in a number of plant species. It comprises a helical region with homology to a region of chromatin binding proteins, an Arg-Gly-Asp cell adhesion motif and eight aspartic acid residues. <it>In vitro </it>studies indicate that lunasin suppresses chemical and oncogene driven transformation of mammalian cells. We have explored efficient recombinant production of lunasin by exploiting the <it>Clostridium thermocellum </it>CipB cellulose binding domain (CBD) as a fusion partner protein.</p> <p>Results</p> <p>We used a pET28 vector to express a CBD-lunasin fusion with a hexahistidine tag and Tobacco Etch Virus protease site, to allow protease-mediated release of native lunasin. Autoinduction in <it>E. coli </it>BL21 (DE3) Star cells achieved expression of 3.35 g/L of CBD-lunasin fusion protein. The final yield of lunasin was 210 mg/L corresponding to 32% of the theoretical yield. Purification by cellulose binding and nickel affinity chromatography were tested with the latter proving more satisfactory.</p> <p>The effects of CBD-lunasin expression on growth and morphology of the <it>E. coli </it>cells were examined by light and electron microscopy revealing an altered morphology in a proportion of cells. Cell division appeared to be inhibited in these cells resulting in elongated, non-septated cells.</p> <p>Conclusions</p> <p>The use of CBD as a fusion partner gave high protein yields by autoinduction, with lunasin release by TEV protease cleavage. With some optimisation this approach could provide a potentially valuable route for production of this therapeutic peptide. Over-expression in the host cells manifest as a cell division defect in a population of the cells, presumably mimicking some aspect of the chemopreventive function observed in mammalian cells.</p

Sports-related brain injury in the general population: An epidemiological study

Objectives To determine the incidence, nature and severity of all sports-related brain injuries in the general population. Design Population-based epidemiological incidence study. Methods Data on all traumatic brain injury events sustained during a sports-related activity were extracted from a dataset of all new traumatic brain injury cases (both fatal and non-fatal), identified over a one-year period in the Hamilton and Waikato districts of New Zealand. Prospective and retrospective case ascertainment methods from multiple sources were used. All age groups and levels of traumatic brain injury severity were included. Details of the registering injuries and recurrent injuries sustained over the subsequent year were obtained through medical/accident records and assessment interviews with participants. Results Of 1369 incident traumatic brain injury cases, 291 were identified as being sustained during a sports-related activity (21% of all traumatic brain injuries) equating to an incidence rate of 170 per 100,000 of the general population. Recurrent injuries occurred more frequently in adults (11%) than children (5%). Of the sports-related injuries 46% were classified as mild with a high risk of complications. Injuries were most frequently sustained during rugby, cycling and equestrian activities. It was revealed that up to 19% of traumatic brain injuries were not recorded in medical notes. Conclusions Given the high incidence of new and recurrent traumatic brain injury and the high risk of complications following injury, further sport specific injury prevention strategies are urgently needed to reduce the impact of traumatic brain injury and facilitate safer engagement in sports activities. The high levels of ‘missed’ traumatic brain injuries, highlights the importance in raising awareness of traumatic brain injury during sports-related activity in the general population

Diversity and Abundance of Club and Coral Fungi in the Upper Lane Cove Valley

The Kingdom Fungi are central players in the ecology and biogeochemistry of terrestrial ecosystems. Despite this importance, the diversity, distribution and abundance of fungal species are poorly known. Here, we undertook an intensive survey of club and coral fungi in the Upper Lane Cove Valley, Sydney, Australia. Over a two-year period, we collected more than 1100 specimens, and identified these to genus using a combination of DNA barcoding and morphology. The majority of specimens did not match any sequences in GenBank at more than 95% similarity, meaning that many of these fungi are either poorly represented in DNA databases, or are potentially novel species. A number of hotspots for fungal diversity and abundance were identified, largely along creek lines draining southwest through coachwood dominated vegetation. Notably, these hotspots all lie outside the adjacent Lane Cove National Park

Community Ideas Factory

The Community Ideas Factory is a community-college partnership exploring social innovations within the charitable sector of the Halton Region. It is a collaborative research project between Sheridan College and the Oakville Community Foundation. The goal of the project is to change the philanthropic granting process in Oakville so that it is more bottom-up, participatory, and evidenced-based. The principle community partner on the project is The Foundation (OCF). A community organization tasked with managing and disbursing donor contributions for philanthropic projects in the Town of Oakville. The Community Ideas Factory is made possible by the College-Community Social Innovation Fund of the Social Sciences and Humanities Research Council of Canada (SSHRC)

Phage display selected magnetite interacting Adhirons for shape controlled nanoparticle synthesis

Adhirons are robust, well expressing, peptide display scaffold proteins, developed as an effective alternative to traditional antibody binding proteins for highly specific molecular recognition applications. This paper reports for the first time the use of these versatile proteins for material binding, and as tools for controlling material synthesis on the nanoscale. A phage library of Adhirons, each displaying two variable binding loops, was screened to identify specific proteins able to interact with [100] faces of cubic magnetite nanoparticles. The selected variable regions display a strong preference for basic residues such as lysine. Molecular dynamics simulations of amino acid adsorption onto a [100] magnetite surface provides a rationale for these interactions, with the lowest adsorption energy observed with lysine. These proteins direct the shape of the forming nanoparticles towards a cubic morphology in room temperature magnetite precipitation reactions, in stark contrast to the high temperature, harsh reaction conditions currently used to produce cubic nanoparticles. These effects demonstrate the utility of the selected Adhirons as novel magnetite mineralization control agents using ambient aqueous conditions. The approach we outline with artificial protein scaffolds has the potential to develop into a toolkit of novel additives for wider nanomaterial fabrication

1004-59 Vascular Acoustic Emissions During Angioplasty: Potential Role in Identification of Induced Dissection

A fundamental mechanism of balloon angioplasty (BA) is plaque rupture. Rupture leading to dissection, however, has been implicated as an underlyIng factor responsible for both acute and chronic adverse outcomes. Acoustic emissions (AE) — transient sound waves generated by microstructural alterations of a material subjected to mechanical stress — may provide a novel means of characterizing BA-induced tissue trauma. Using a novel acoustic sensor system, we examined the relationship between cumulative AE energy released by human arterial tissue during BA and the observed pathologic injury. Post-mortem human arterial specimens (19) were subjected to identical SA with simultaneous monitoring of intraluminal pressure and AE. Sound energy was integrated throughout the pressurization period to obtain an estimate of the cumulative AE energy released during dilatation. Postangioplasty inspection revealed a marked difference in AE energy released by specimens that experienced traumatic dissection vs. non-dissection dilatation:Sound energy released by vascular tissue undergoing balloon angioplasty discriminates dissection from non-dissection tissue trauma. Given the deleterious role that dissection can play in SA, this novel system may provide a means of improving procedural outcome

Band gap modulation of zirconium-based metal-organic frameworks by defect engineering

We report a defect-engineering approach to modulate the band gap of zirconium-based metal-organic framework UiO-66, enabled by grafting of a range of amino-functionalised benzoic acids at defective sites. Defect engineered MOFs were obtained by both post-synthetic exchange and modulated synthesis, featuring band gap in the 4.1-3.3 eV range. First principle calculations suggest that shrinking of the band gap is likely due to an upward shift of the valence band energy, as a result of the presence of light-absorbing monocarboxylates. The photocatalytic properties of defect-engineered MOFs towards CO2 reduction to CO in the gas phase and degradation of Rhodamine B in water were tested, observing improved activity in both cases, in comparison to a defective UiO-66 bearing formic acid as the defect-compensating species

Nicked-site substrates for a serine recombinase reveal enzyme-DNA communications and an essential tethering role of covalent enzyme-DNA linkages

To analyse the mechanism and kinetics of DNA strand cleavages catalysed by the serine recombinase Tn3 resolvase, we made modified recombination sites with a single-strand nick in one of the two DNA strands. Resolvase acting on these sites cleaves the intact strand very rapidly, giving an abnormal half-site product which accumulates. We propose that these reactions mimic second-strand cleavage of an unmodified site. Cleavage occurs in a synapse of two sites, held together by a resolvase tetramer; cleavage at one site stimulates cleavage at the partner site. After cleavage of a nicked-site substrate, the half-site that is not covalently linked to a resolvase subunit dissociates rapidly from the synapse, destabilizing the entire complex. The covalent resolvase–DNA linkages in the natural reaction intermediate thus perform an essential DNA-tethering function. Chemical modifications of a nicked-site substrate at the positions of the scissile phosphodiesters result in abolition or inhibition of resolvase-mediated cleavage and effects on resolvase binding and synapsis, providing insight into the serine recombinase catalytic mechanism and how resolvase interacts with the substrate DNA

Emergence of good conduct, scaling and Zipf laws in human behavioral sequences in an online world

We study behavioral action sequences of players in a massive multiplayer online game. In their virtual life players use eight basic actions which allow them to interact with each other. These actions are communication, trade, establishing or breaking friendships and enmities, attack, and punishment. We measure the probabilities for these actions conditional on previous taken and received actions and find a dramatic increase of negative behavior immediately after receiving negative actions. Similarly, positive behavior is intensified by receiving positive actions. We observe a tendency towards anti-persistence in communication sequences. Classifying actions as positive (good) and negative (bad) allows us to define binary 'world lines' of lives of individuals. Positive and negative actions are persistent and occur in clusters, indicated by large scaling exponents alpha~0.87 of the mean square displacement of the world lines. For all eight action types we find strong signs for high levels of repetitiveness, especially for negative actions. We partition behavioral sequences into segments of length n (behavioral `words' and 'motifs') and study their statistical properties. We find two approximate power laws in the word ranking distribution, one with an exponent of kappa-1 for the ranks up to 100, and another with a lower exponent for higher ranks. The Shannon n-tuple redundancy yields large values and increases in terms of word length, further underscoring the non-trivial statistical properties of behavioral sequences. On the collective, societal level the timeseries of particular actions per day can be understood by a simple mean-reverting log-normal model.Comment: 6 pages, 5 figure