Analysis of UDP-galactose 4'-epimerase mutations associated with the intermediate form of type III galactosaemia

Type III galactosaemia is a hereditary disease caused by reduced activity in the Leloir pathway enzyme, UDP-galactose 4'-epimerase (GALE). Traditionally, the condition has been divided into two forms-a mild, or peripheral, form and a severe, or generalized, form. Recently it has become apparent that there are disease states which are intermediate between these two extremes. Three mutations associated with this intermediate form (S81R, T150M and P293L) were analysed for their kinetic and structural properties in vitro and their effects on galactose-sensitivity of Saccharomyces cerevisiae cells that were deleted for the yeast GALE homologue Gal10p. All three mutations result in impairment of the kinetic parameters (principally the turnover number, k(cat)) compared with the wild-type enzyme. However, the degree of impairment was mild compared with that seen with the mutation (V94M) associated with the generalized form of epimerase deficiency galactosaemia. None of the three mutations tested affected the ability of the protein to dimerize in solution or its susceptibility to limited proteolysis in vitro. Finally, in the yeast model, each of the mutated patient alleles was able to complement the galactose-sensitivity of gal10 Delta cells as fully as was the wild-type human allele. Furthermore, there was no difference from control in metabolite profile following galactose exposure for any of these strains. Thus we conclude that the subtle biochemical and metabolic abnormalities detected in patients expressing these GALE alleles likely reflect, at least in part, the reduced enzymatic activity of the encoded GALE proteins.

The distribution of Fasciola hepatica and Fasciola gigantica within southern Tanzania - constraints associated with the intermediate host

In East Africa, Fasciola gigantica is generally the causative agent of fasciolosis but there have been reports of F. hepatica in cattle from highland regions of Kenya, Ethiopia, Uganda and Zaire. The topography of the Southern Highlands of Tanzania provides an environment where the climatic conditions exist for the sustenance of lymnaeid species capable of Supporting both Fasciola hepatica and F. gigantica. Theoretically this would allow interaction between fasciolid species and the possible creation of hybrids. In this report we present molecular data confirming the existence of the snail, Lymnaea truncatula, at high altitude on the Kitulo Plateau of the Southern Highlands, Tanzania, along with morphometric and molecular data confirming the presence of F. hepatica in the corresponding area. At lower altitudes, where climatic conditions were unfavourable for the existence of L. truncatula, the presence of its sister species L. natalensis was confirmed by molecular data along with its preferred fasciolid parasite, F. gigantica. Analysis based on a 618 bp sequence of the 28S rRNA gene did not reveal the presence of hybrid fasciolids in our fluke samples.

Extraction of electrode kinetic parameters from microdisc voltammetric data measured under transport conditions intermediate between steady-state convergent and transient linear diffusion as typically applies to room temperature ionic liquids

The extraction of electrode kinetic parameters for electrochemical couples in room-temperature ionic liquids (RTILs) is currently an area of considerable interest. Electrochemists typically measure electrode kinetics in the limits of either transient planar or steady-state convergent diffusion for which the voltammetic response is well understood. In this paper we develop a general method allowing the extraction of this kinetic data in the region where the diffusion is intermediate between the planar and convergent limits, such as is often encountered in RTILs using microelectrode voltammetry. A general working surface is derived, allowing the inference of Butler-Volmer standard electrochemical rate constants for the peak-to-peak potential separation in a cyclic voltammogram as a function of voltage scan rate. The method is applied to the case of the ferrocene/ferrocenium couple in [C(2)mim][N(Tf)(2)] and [C(4)mim][N(Tf)(2)].

A density functional theory study of hydroxyl and the intermediate in the water formation reaction on Pt

Density functional theory has been used to study the adsorption of hydroxyl at low and high coverages and also to investigate the nature of the intermediate in the H2O formation reaction on Pt(111). At low coverages [1/9 of a monolayer (ML) to 1/3 ML] OH binds preferentially at bridge and top sites with a chemisorption energy of similar to2.25 eV. At high coverages (1/2 ML to 1 ML) H bonding between adjacent hydroxyls causes: (i) an enhancement in OH chemisorption energy by about 15%; (ii) a strong preference for OH adsorption at top sites; and (iii) the formation of OH networks. The activation energy for the diffusion of isolated OH groups along close packed rows of Pt atoms is 0.1 eV. This low barrier coupled with H bonding between neighboring OH groups indicates that hydroxyls are susceptible to island formation at low coverages. Pure OH as well as coadsorbed OH and H can be ruled out as the observed low temperature intermediate in the water formation reaction. Instead we suggest that the intermediate consists of a mixed OH+H2O overlayer with a macroscopic surface coverage of 3/4 ML in a 2:1 ratio of OH and H2O. (C) 2001 American Institute of Physics.

Observations towards early-type stars in the ESO-POP Survey - II. Searches for intermediate- and high-velocity clouds

We present Ca it K and Ti it optical spectra of early-type stars taken mainly from the ultraviolet and visual echelle spectrograph (LIVES) Paranal Observatory Project, plus H 1 21-cm spectra, from the Vila-Elisa and Leiden-Dwingeloo Surveys, which are employed to obtain distances to intermediate- and high-velocity clouds (IHVCs). H I emission at a velocity of -117 km s(-1) towards the sightline HD 30677 (l, b = 190 degrees.2, -22 degrees.2) with column density -1.7 x 10(19) cm(-2) has no corresponding Ca Pi K absorption in the LIVES spectrum, which has a signal-to-noise ratio (S/N) of 610 per resolution element. The star has a spectroscopically determined distance of 2.7 kpc, and hence sets this as a firm lower distance limit towards Anti-Centre cloud ACII. Towards another sightline (HD 46185 with 1, b = 222 0, -10 degrees.1), H1 at a velocity of +122 km s(-1) and column density of 1.2 x 10(19) cm(-2) is seen. The corresponding Ca Pi K spectrum has a S/N of 780, although no absorption is observed at the cloud velocity. This similarly places a firm lower distance limit of 2.9 kpc towards this parcel of gas that may be an intermediate-velocity (IV) cloud. The lack of IV Ca it absorption towards HD 196426 (1, b = 45 degrees.8, -23 degrees.3) at a S/N of 500 reinforces a lower distance limit of -700 pc towards this part of complex gp, where the H I column density is 1.1 x 1019 cm(-2) and velocity is +78 km s(-1). Additionally, no IV Cart is seen in absorption in the spectrum of HD 19445, which is strong in H I with a column density of 8 x 10(19) cm(-2) at a velocity of - -42 km s(-1), placing a firm although uninteresting lower distance limit of 39 pc to this part of IV South. Finally, no high-velocity Call K absorption is seen towards HD 115363 (l, b = 306.0,-1.0) at a S/N of 410, placing a lower distance of -3.2 kpc towards the HVC gas at velocity of - +224 km s(-1) and WE column density of 5.2 x 10(19) cm(-2). This gas is in the same region of the sky as complex WE (Wakker 2001), but at higher velocities. The non-detection of Ca it K absorption sets a lower distance of -3.2 kpc towards the HVC, which is unsurprising if this feature is indeed related to the Magellanic System.

Child abuse and neglect: Training needs of student teachers

Increasing awareness of child abuse and neglect (CAN) raises questions about how well teachers are prepared for their role in child protection. This paper assesses and differentiates training needs of first-year students (n = 216) in Northern Ireland. Multiplechoice tests were used to assess knowledge of CAN statistics; recognising and reporting; policies, procedures, and legislative frameworks; and direct work with children. Considerable gaps in knowledge were found. Results between student groups varied and provide evidence of the need to develop pre-service child protection training. The importance of differentiation between student groups in terms of training content is emphasised.

Palladium catalysed formal 6-endo-trig approaches to pumiliotoxin alkaloids: interception of the elusive cyclopropyl intermediate

Intramolecular Heck cyclisation of (E)-vinyl bromides leads to indolizidines, related to pumiliotoxin alkaloids, in which the stereochemistry of the trisubstituted double bond undergoes inversion. A cyclopropyl intermediate, which is believed to be responsible for the double bond inversion, has been intercepted by forcing an 'early' beta-hydride elimination on this species. The relative stereochemistry of this cyclopropyl intermediate determines the regioselectivity of the final beta-hydride elimination. In this case all three beta-hydride eliminations were stereochemically permitted, giving rise to a mixture of three isomeric products, differing in the position of a double bond. (Z)-Vinyl bromides were found to be less reactive than (E)-vinyl bromides, but on cyclisation gave the required conjugated diene, with inversion of the vinyl bromide stereochemistry, as the sole reaction product. This methodology will allow rapid stereoselective access to the diene-based pumiliotoxin alkaloids.