Production, crystallization and preliminary X-ray analysis of CTP: inositol-1-phosphate cytidylyltransferase from Archaeoglobus fulgidus

Brito, J. A.Archaeoglobus fulgidus, a hyperthermophilic archaeon, accumulates di-myo-inositol phosphate (DIP) in response to heat stress. Recently, the pathway for biosynthesis of DIP has been elucidated in this organism and involves a bifunctional enzyme that contains two domains: CTP:inositol-1-phosphate cytidylyltransferase (IPCT) as a soluble domain and di-myo-inositol-1,3'-phosphate-1-phosphate synthase (DIPPS) as a membrane domain. Here, the expression, purification, crystallization and preliminary X-ray diffraction analysis of the IPCT domain from A. fulgidus in the apo form are reported. The crystals diffracted to 2.4 A resolution using a synchrotron source and belonged to the orthorhombic space group P2(1)2(1)2, with unit-cell parameters a = 154.7, b = 83.9, c = 127.7 A.publishe

Molecular epidemiology of Streptococcus uberis clinical mastitis in dairy herds: strain heterogeneity and transmission

Multi Locus Sequence Typing was successfully completed on 494 isolates of S. uberis from clinical mastitis cases in a study of 52 commercial dairy herds over a 12 month period. In total, 195 sequence types (STs) were identified. S. uberis mastitis cases occurring in different cows within the same herd and attributed to a common ST were classified as 'potential transmission events' (PTE). Clinical cases attributed to 35 of the 195 STs identified in this study were classified PTE. PTE were identified in 63% of herds. PTE associated cases, which include the first recorded occurrence of that ST in that herd (Index case) and all persistent infections with that PTE ST represented 40% of all the clinical mastitis cases and occurred in 63% of herds. PTE associated cases accounted for over 50% of all S. uberis clinical mastitis cases in 33% of herds. Nine sequence types (ST 5, 6, 20, 22, 24, 35, 233, 361, and 512), eight of which grouped within a clonal complex (sharing at least four alleles), were statistically overrepresented (OVR STs). The findings indicate that 38% of all clinical mastitis cases and 63% of the potential transmission events attributed to S. uberis in dairy herds may be caused by the nine most prevalent strains. The findings suggest that to a small subset of sequence types are disproportionally important in the epidemiology of S. uberis mastitis in the UK with cow to cow transmission of S. uberis potentially occurring in the majority of UK herds and may be the most important route of infection in many herds

Fatty Acid Methyl Esters as Biosolvents of Epoxy Resins: A Physicochemical Study

The C8 to C18 fatty acid methyl esters (FAME) have been compared as solvents for two epoxy resin pre-polymers, bisphenol A diglycidyl ether (DGEBA) and triglycidyl paminophenol ether (TGPA). It was found that the solubilization limits vary according to the ester and that methyl caprylate is the best solvent of both resins. To explain these solubility performances, physical and chemical properties of FAME were studied, such as the Hansen parameters, viscosity, binary diffusion coefficient and vaporization enthalpy. Determination of the physicochemical parameters of FAME was carried out by laboratory experimentations and by calculation from bibliographic data. The Hansen parameters of FAME and epoxy resins pre-polymers were theoretically and experimentally determined. The FAME chain length showed a long dependence on the binary diffusion parameters and kinematic viscosity, which are mass and momentum transport properties. Moreover, the vaporization enthalpy of these compounds was directly correlated with the solubilization limits

Homo- and Heteroleptic Phototoxic Dinuclear Metallo-Intercalators Based on Ru II (dppn) Intercalating Moieties: Synthesis, Optical and Biological Studies

Using a new mononuclear “building block,” for the first time, a dinuclear RuII(dppn) complex and a heteroleptic system containing both RuII(dppz) and RuII(dppn) moieties are reported. The complexes, including the mixed dppz/dppn system, are 1O2 sensitizers. However, unlike the homoleptic dppn systems, the mixed dppz/dppn complex also displays a luminescence “switch on” DNA light-switch effect. In both cisplatin sensitive and resistant human ovarian carcinoma lines the dinuclear complexes show enhanced uptake compared to their mononuclear analogue. Thanks to a favorable combination of singlet oxygen generation and cellular uptake properties all three of the new complexes are phototoxic and display potent activity against chemotherapeutically resistant cells

Educational change in Scotland: Policy, context and biography

The poor success rate of policy for curriculum change has been widely noted in the educational change literature. Part of the problem lies in the complexity of schools, as policymakers have proven unable to micromanage the multifarious range of factors that impact upon the implementation of policy. This paper draws upon empirical data from a local authority-led initiative to implement Scotland’s new national curriculum. It offers a set of conceptual tools derived from critical realism (particularly the work of Margaret Archer), which offer significant potential in allowing us to develop greater understanding of the complexities of educational change. Archer’s social theory developed as a means of explaining change and continuity in social settings. As schools and other educational institutions are complex social organisations, critical realism offers us epistemological tools for tracking the ebbs and flows of change cycles over time, presenting the means for mapping the multifarious networks and assemblages that form their basis

A dinuclear ruthenium(II) complex excited by near-infrared light through two-photon absorption induces phototoxicity deep within hypoxic regions of melanoma cancer spheroids

The dinuclear photo-oxidizing RuII complex [{Ru(TAP2)}2(tpphz)]4+ (TAP = 1,4,5,8- tetraazaphenanthrene, tpphz = tetrapyrido[3,2-a:2',3'-c:3'',2''- h:2''',3'''-j]phenazine), 14+ is readily taken up by live cells localizing in mitochondria and nuclei. In this study, the two-photon absorption cross-section of 14+ is quantified and its use as a two-photon absorbing phototherapeutic is reported. It was con-firmed that the complex is readily photo-excited using near infrared, NIR, light through two-photon absorption, TPA. In 2-D cell cul-tures, irradiation with NIR light at low power results in precisely focused photo-toxicity effects in which human melanoma cells were killed after 5 minutes of light exposure. Similar experiments were then carried out in human cancer spheroidsthat provide a realistic tumor model for the development of therapeutics and phototherapeutics. Using the characteristic emission of the complex as a probe, its up-take into 280 µm spheroids was investigated and confirmed that the spheroid takes up the complex. Notably TPA excitation results in more intense luminescence being observed throughout the depth of the spheroids, although emission intensity still drops off toward the necrotic core. As 14+ can directly photo-oxidize DNA without the mediation of singlet oxygen or other reactive oxygen species, photo-toxicity within the deeper, hypoxic layers of the spheroids was also investigated. To quantify the penetration of these phototoxic effects, 14+ was photo-excited through TPA at a power of 60 mW, which was progressively focused in 10 µm steps throughout the entire z-axis of individual spheroids. These experiments revealed that, in irradiated spheroids treated with 14+, acute and rapid photo-induced cell death was observed throughout their depth, including the hypoxic region

Making the right link to theranostics : the photophysical and biological properties of dinuclear Ru^II-Re^I dppz complexes depend on their tether

The synthesis of new dinuclear complexes containing linked RuII(dppz) and ReI(dppz) moieties is reported. The photophysical and biological properties of the new complex, which incorporates a N,N′-bis(4-pyridylmethyl)-1,6-hexanediamine tether ligand, are compared to a previously reported RuII/ReI complex linked by a simple dipyridyl alkane ligand. Although both complexes bind to DNA with similar affinities, steady-state and time-resolved photophysical studies reveal that the nature of the linker affects the excited state dynamics of the complexes and their DNA photocleavage properties. Quantum-based DFT calculations on these systems offer insights into these effects. While both complexes are live cells permeant, their intracellular localizations are significantly affected by the nature of the linker. Notably, one of the complexes displayed concentration-dependent localization and possesses photophysical properties that are compatible with SIM and STED nanoscopy. This allowed the dynamics of its intracellular localization to be tracked at super resolutions