Biomedical subjects

Peter A Williams

Publications and source records attributed to Peter A Williams.

At least 19 recordsLinked to original sources

PubMed2006

Solution properties of targacanthin (water-soluble part of gum tragacanth exudate from Astragalus gossypinus).

Solution properties of tragacanthin (the water-soluble part of gum tragacanth) were studied by gel permeation chromatography (GPC) combined with multi-angle light scattering and viscometry at 25 degrees C. Photon correlation spectroscopy was used to determine the hydrodynamic radius. Ultrasonic degradation was applied to obtain biopolymer fractions of different molecular weights. The dependence of intrinsic viscosity [eta] and radius of gyration (s2)z(1/2) on weight average molecular mass M(w) for this biopolymer were found to be [eta] = 9.077 x 10(-5) M(w)(0.87) (dL g(-1)) and (s2)z(1/2) in the range of M(w) from 1.8 x 10(5) to 1.6 x 10(6). The conformational parameters of tragacanthin were calculated to be 1111 nm for molar mass per unit contour length (M(L)), 26 nm for persistence length (q) and 1.87 ratio of R(g)/R(h). It was found that the Smidsrød parameter B, the empirical stiffness parameter was 0.013, which is lower than that of several polysaccharides indicating the stiff backbone for tragacanthin. The rheological behavior of aqueous solutions of gum tragacanth and its insoluble and soluble fractions (bassorin and tragacanthin, respectively) were studied. For concentrations equal to 1%, at 25 degrees C and in the absence of salt, bassorin solution showed the highest viscosity and shear thinning behaviour. Power law and Williamson models were used to describe the rheological behaviour of bassorin and tragacanthin, respectively. Oscillatory shear experiments showed a gel like structure for the bassorin but for tragacanthin the oscillatory data were as would be expected for semi-dilute to concentrated solution of entangled, random coil polymers. NaCl changed the steady and oscillatory rheological properties of both fractions and in this way the final viscosity of bassorin was even less than tragacanthin. The calculated activation energy for bassorin and tragacanthin indicated a more rapid decrease in viscosity with temperature for tragacanthin. The plot of eta(sp,0) versus C[eta] revealed that the transition from dilute to semi-dilute regime occurs at C*[eta] = 2.82 for tragacanthin.

Astragalus Plant↗

PubMed2006

Hybrid density functional theory for pi-stacking interactions: application to benzenes, pyridines, and DNA bases.

The suitability of a hybrid density functional to qualitatively reproduce geometric and energetic details of parallel pi-stacked aromatic complexes is presented. The hybrid functional includes an ad hoc mixture of half the exact (HF) exchange with half of the uniform electron gas exchange, plus Lee, Yang, and Parr's expression for correlation energy. This functional, in combination with polarized, diffuse basis sets, gives a binding energy for the parallel-displaced benzene dimer in good agreement with the best available high-level calculations reported in the literature, and qualitatively reproduces the local MP2 potential energy surface of the parallel-displaced benzene dimer. This method was further critically compared to high-level calculations recently reported in the literature for a range of pi-stacked complexes, including monosubstituted benzene-benzene dimers, along with DNA and RNA bases, and generally agrees with MP2 and/or CCSD(T) results to within +/-2 kJ mol(-1). We also show that the resulting BH&H binding energy is closely related to the electron density in the intermolecular region. The net result is that the BH&H functional, presumably due to fortuitous cancellation of errors, provides a pragmatic, computationally efficient quantum mechanical tool for the study of large pi-stacked systems such as DNA.

Algorithms↗

PubMed2005

Raman spectroscopy of likasite at 298 and 77 K.

Raman spectroscopy at 298 and 77K has been used to study the structure of likasite, a naturally occurring basic copper(II) nitrate of formula Cu3NO3(OH)5.2H2O. An intense sharp band is observed at 3522 cm(-1) at 298 K which splits into two bands at 3522 and 3505 cm(-1) at 77 K and is assigned to the OH stretching mode. The two OH stretching bands at 3522 and 3505 provide estimates of the hydrogen bond distances of these units as 2.9315 and 2.9028 angstroms. The significance of this result is that equivalent OH units in the 298 K spectrum become two non-equivalent OH units at 77 K suggesting a structural change by cooling to liquid nitrogen temperature. A number of broad bands are observed in the 298 K spectrum at 3452, 3338, 3281 and 3040 cm(-1) assigned to H2O stretching vibrations with estimates of the hydrogen bond distances of 2.8231, 2.7639, 2.7358 and 2.6436 angstroms. Three sharp bands are observed at 77 K at 1052, 1050 and 1048 cm(-1) attributed to the nu1 symmetric stretching mode of the NO3 units. Only a single band at 1050 cm(-1) is observed at 298 K, suggesting the non-equivalence of the NO3 units at 77 K, confirming structural changes in likasite by cooling to 77 K.

Chemistry, Physical↗

PubMed2004

The development of immunoassays to identify and quantify species source of gum arabic.

Gum arabic from Acacia senegal is commonly used as an additive in foodstuffs. Adulteration of gum arabic by other gums is a potential problem for reasons of safety and quality. This study aimed to develop and evaluate the use of enzyme-linked immunosorbent assays (ELISAs) for the detection of potential adulterants of gum arabic. Indirect competitive ELISAs (IC-ELISAs) were developed using the monoclonal antibodies SY CC7 (A. senegal), SY HH3 (Acacia seyal), and SY J1A1 (Combretum erythrophyllum). All IC-ELISAs had a working range of 0.005-10 mg/mL. The antibodies used were tested using the IC-ELISAs for cross-reactivity with other Acacia species and other gums. The antibodies were very specific for their respective antigens. Significant cross-reactivity was found for SY CC7 (between A. senegal and A. melliferae) and SY J1A1 (between C. erythrophyllum and A. seyal). The IC-ELISA was adapted further to test confectionery samples for the presence of gum arabic, which was successful, although recovery rates were reduced. Both IC- and plate trapped antigen ELISA (PTA-ELISA) formats were able to distinguish an adulterated sample of gum arabic when blended with either A. seyal or C. erythrophyllum. The PTA-ELISA was more sensitive for A. seyal than the IC-ELISA, but both were equally sensitive for C. erythrophyllum. The results suggest that the antibodies SY CC7, SY HH3, and SY J1A1 could be used in combination with each other for the detection of potential adulterants of A. senegal and the detection of gum arabic in foodstuffs.

Acacia↗

PubMed2004

Raman spectroscopy of some basic chloride containing minerals of lead and copper.

Raman spectroscopy has been used to characterise several lead and mixed cationic-lead minerals including mendipite, perite, laurionite, diaboléite, boléite, pseudoboléite, chloroxiphite, and cumengéite. Raman spectroscopy enables their vibrational spectra to be compared. The low wavenumber region is characterised by the bands assigned to cation-chloride stretching and bending modes. Phosgenite is a mixed chloride-carbonate mineral and a comparison is made with the molecular structure of the aforementioned minerals. Each mineral shows different hydroxyl stretching vibrational patterns, but some similarity exists in the Raman spectra of the hydroxyl deformation modes. Raman spectroscopy lends itself to the study of these types of minerals in complex mineral systems involving secondary mineral formation.

Chlorides↗

PubMed2004

Use of infrared spectroscopy for the determination of electronegativity of rare earth elements.

Infrared spectroscopy has been used to study a series of synthetic agardite minerals. Four OH stretching bands are observed at around 3568, 3482, 3362, and 3296 cm(-1). The first band is assigned to zeolitic, non-hydrogen-bonded water. The band at 3296 cm(-1) is assigned to strongly hydrogen-bonded water with an H bond distance of 2.72 A. The water in agardites is better described as structured water and not as zeolitic water. Two bands at around 999 and 975 cm(-1) are assigned to OH deformation modes. Two sets of AsO symmetric stretching vibrations were found and assigned to the vibrational modes of AsO(4) and HAsO(4) units. Linear relationships between positions of infrared bands associated with bonding to the OH units and the electronegativity of the rare earth elements were derived, with correlation coefficients >0.92. These linear functions were then used to calculate the electronegativity of Eu, for which a value of 1.1808 on the Pauling scale was found.

Journal Article↗

PubMed2003

Role of hydrocolloids in the creaming of oil in water emulsions.

The influence of guar gum and xanthan gum on the creaming of 10% oil in water emulsions has been investigated. The presence of very low concentrations of the polysaccharides (typically < approximately 0.075%) was found to induce depletion flocculation of the emulsion droplets and increased the rate of creaming. However, at higher polysaccharide concentrations (typically > approximately 0.1%) the creaming rate was reduced due to the increased viscosity of the aqueous phase. For most systems a delay period was noted before creaming started, which was found to be dependent on the zero shear viscosity of the continuous phase and independent of polysaccharide type. The delay period increased significantly at zero shear viscosities approaching 1 Pa s. A mathematical model has been used to fit the creaming rate profiles and a simple exponential relationship obtained between delay time and polysaccharide concentration.

Chemical Phenomena↗

PubMed2003

Infrared spectroscopic study of natural hydrotalcites carrboydite and hydrohonessite.

Infrared spectroscopy has proven most useful for the study of anions in the interlayer of natural hydrotalcites. A suite of naturally occurring hydrotalcites including carrboydite, hydrohonessite, reevesite, motukoreaite and takovite were analysed. Variation in the hydroxyl stretching region was observed and the band profile is a continuum of states resulting from the OH stretching of the hydroxyl and water units. Infrared spectroscopy identifies some isomorphic substitution of sulphate for carbonate through an anion exchange mechanism for the minerals carrboydite and hydrohonessite. The infrared spectra of the CO3 and SO4 stretching region of takovite is complex because of band overlap. For this mineral some sulphate has replaced the carbonate in the structure. In the spectra of takovites, a band is observed at 1346 cm(-1) and is attributed to the carbonate anion hydrogen bonded to water in the interlayer. Infrared spectroscopy has proven most useful for the study of the interlayer structure of these natural hydrotalcites.

Aluminum Hydroxide↗

PubMed2003

Molecular weight, tertiary structure, water binding and colon behaviour of ispaghula husk fibre.

Molecular variables, using aqueous and alkaline extracts, of the polysaccharide from ispaghula husk (IH) were examined using gel-permeation chromatography linked to multi-angle laser light scattering. Progressive extraction can yield a component with a molecular weight (MW)value up to about 7 x 106 Da, and gels, which accompany the extraction, have MW ranging from 10-20 x 106 Da. To mimic the polysaccharide degradation, particularly in the colon, the solid IH was degraded progressively using ionising radiation. A chain break occurs every 7.5 kGy in NaOH and every 15 kGy in water. The solid-state matrix is opened by the radiation to yield increased visco-elasticity of the aqueous extracts at critical radiation doses, before further degradation occurs after about 12 kGy. Differential scanning calorimetry is used to study the mechanism of interaction of water with IH. The first water to be taken up is non-freezing water and represents about twelve water molecules/disaccharide unit of the polysaccharide. As the water content is increased, the water becomes bound to the polysaccharide and freezes and melts at a temperature different from free water. This water is thermodynamically distinguishable from free water. It forms amorphous ice on cooling which crystallises exothermically and subsequently melts endothermically. Saturation occurs at a water content of 2-3 g water/g polymer, showing that about 60% of the water in the system is 'bound'. The most surprising conclusion is that despite the fact that the IH swells in water to form a solid and stiff gel, the greater part of that water in the gel is still free and behaves like liquid water.

Chromatography, Gel↗

PubMed2003

Mutational analysis of the critical bases involved in activation of the AreR-regulated sigma54-dependent promoter in Acinetobacter sp. strain ADP1.

The areR gene in Acinetobacter sp. strain ADP1 regulates the expression of the areCBA genes, which determine growth on benzyl alkanoates. AreR is a member of the NtrC/XylR family of regulatory proteins as determined by sequence homology. Seventy-nine bases upstream of the start of transcription is a region carrying two overlapping inverted repeat (IR) sequences that we predict to be the AreR binding site, also known as the upstream activator site (UAS). IR1 is a near-perfect (16 of 17 bp) repeat separated by 1 bp, and IR2 consists of 9- and 7-bp perfect repeats with a 3-bp gap, with the central bases of the two arms of the repeat separated by 44 and 22 bp. We report here a method for site-directed mutagenesis of chromosomal genes in ADP1 in which linear fragments generated by overlap extension PCR are used to transform ADP1 via its natural transformation system and recombinants are selected by a marker exchange-eviction strategy with a newly created sacB-Km cassette. This method was used to generate 38 strains with designed mutations in the putative UAS upstream of areCBA. The effects of the mutations on areCBA expression were measured by enzyme assays of benzyl alcohol dehydrogenase (AreB) and by reporter gene assays of lacZ inserted into areA. Substitutions or deletions in IR1 had more deleterious effects upon expression when they were in its central region, which overlaps the left arm of IR2, than when they were in its outer regions. By contrast, substitutions in the right arm of IR2 resulted in mutants with relatively high expression levels compared to that of the wild type. Effects of deletions in the right arm of IR2 were very dependent upon the length of the deletion, with 3- or 5-bp deletions reducing expression by >90% whereas an 11-bp deletion in the same area reduced the expression levels by only 50%, suggesting that alterations in the distance and the orientation of the UAS relative to the -24, -12 sigma(54) promoter are critical.

Actinobacteria↗

PubMed2003

The naphthalene catabolic (nag) genes of Ralstonia sp. strain U2 are an operon that is regulated by NagR, a LysR-type transcriptional regulator.

In Ralstonia sp. strain U2, the nag catabolic genes, which encode the enzymes for the pathway that catabolizes naphthalene via the alternative ring cleavage gentisate pathway, are transcribed as an operon under the same promoter. nagR, which encodes a LysR-type transcriptional regulator, is divergently transcribed compared to the nag catabolic genes. A 4-bp frameshift deletion in nagR demonstrated that NagR is required for expression of the nag operon. The transcriptional start of the nag operon was mapped, and a putative -10, -35 sigma(70)-type promoter binding site was identified. Further upstream, a site proximal to the promoter was identified as a site that has bases which have been found to be conserved in the activator-binding motif of other naphthalene pathways. Transcriptional fusion studies demonstrated that NagR regulates the expression of the nag operon positively in the presence of salicylate and to a lesser extent in the presence of 2-nitrobenzoate. Mutation of the LysR-type activator-binding motif in the nag promoter-proximal region resulted in a loss of inducibility of a lacZ reporter gene transcriptionally fused to nagAa, the first gene of the operon. However, other mutations in the region increased the effectiveness of salicylate as an inducer.

Bacterial Proteins↗

PubMed2002

Ecology: Darwin's naturalization hypothesis challenged.

Naturalized plants can have a significant ecological and economic impact, yet they comprise only a fraction ot the plant species introduced by humans. Darwin proposed that introduced plant species will be less likely to establish a self-sustaining wild population in places with congeneric native species because the introduced plants have to compete with their close relatives, or are more likely to be attacked by native herbivores or pathogens, a theory known as Darwin's naturalization hypothesis. Here we analyze a complete list of seed-plant species that have been introduced to New Zealand and find that those with congeneric relatives are significantly more, not less, likely to naturalize--perhaps because they share with their native relatives traits that pre-adapt them to their new environment.

Adaptation, Physiological↗

PubMed2002

Vibrational spectroscopy of the basic copper phosphate minerals: pseudomalachite, ludjibaite and reichenbachite.

The vibrational spectra of pseudomalachite, reichenbachite and ludjibaite have been obtained at 298 K using a combination of FTIR and Raman microscopy. The vibrational spectra of the minerals are different, in line with differences in crystal structure and composition. Some similarity in the Raman spectra of the three polymorphs pseudomalachite, reichenbachite and ludjibaite exists, particularly in the OH stretching region, but characteristic differences in the OH deformation regions are observed. Differences are also observed in the phosphate stretching and deformation regions.

Copper↗

PubMed2002

Complete sequence of the IncP-9 TOL plasmid pWW0 from Pseudomonas putida.

The TOL plasmid pWW0 (117 kb) is the best studied catabolic plasmid and the archetype of the IncP-9 plasmid incompatibility group from Pseudomonas. It carries the degradative (xyl) genes for toluenes and xylenes within catabolic transposons Tn4651 and Tn4653. Analysis of the complete pWW0 nucleotide sequence revealed 148 putative open reading frames. Of these, 77 showed similarity to published sequences in the available databases predicting functions for: plasmid replication, stable maintenance and transfer; phenotypic determinants; gene regulation and expression; and transposition. All identifiable transposition functions lay within the boundaries of the 70 kb transposon Tn4653, leaving a 46 kb sector containing all the IncP-9 core functions. The replicon and stable inheritance region was very similar to the mini-replicon from IncP-9 antibiotic resistance plasmid pM3, with their Rep proteins forming a novel group of initiation proteins. pWW0 transfer functions exist as two blocks encoding putative DNA processing and mating pair formation genes, with organizational and sequence similarity to IncW plasmids. In addition to the known Tn4651 and IS1246 elements, two additional transposable elements were identified as well as several putative transposition functions, which are probably genetic remnants from previous transposition events. Genes likely to be responsible for known resistance to ultraviolet light and free radicals were identified. Other putative phenotypic functions identified included resistance to mercury and other metal ions, as well as to quaternary ammonium compounds. The complexity and size of pWW0 is largely the result of the mosaic organization of the transposable elements that it carries, rather than the backbone functions of IncP-9 plasmids.

Bacterial Proteins↗

PubMed2002

Accumulation of 2-aminophenoxazin-3-one-7-carboxylate during growth of Pseudomonas putida TW3 on 4-nitro-substituted substrates requires 4-hydroxylaminobenzoate lyase (PnbB).

During growth of Pseudomonas putida strain TW3 on 4-nitrotoluene (4NT) or its metabolite 4-nitrobenzoate (4NB), the culture medium gradually becomes yellow-orange with a lambda(max) of 446 nm. The compound producing this color has been isolated and identified as a new phenoxazinone, 2-aminophenoxazin-3-one-7-carboxylate (APOC). This compound is formed more rapidly and in greater quantity when 4-amino-3-hydroxybenzoate (4A3HB) is added to growing cultures of strain TW3 and is also formed nonbiologically when 4A3HB is shaken in mineral salts medium but not in distilled water. It is postulated that APOC is formed by the oxidative dimerization of 4A3HB, although 4A3HB has not been reported to be a metabolite of 4NT or a product of 4NB catabolism by strain TW3. Using the cloned pnb structural genes from TW3, we demonstrated that the formation of the phenoxazinone requires 4-hydroxylaminobenzoate lyase (PnbB) activity, which converts 4-hydroxylaminobenzoate (4HAB) to 3,4-dihydroxybenzoate (protocatechuate) and that 4-nitrobenzoate reductase (PnbA) activity, which causes the accumulation of 4HAB from 4NB, does not on its own result in the formation of APOC. This rules out the possibility that 4A3HB is formed abiotically from 4HAB by a Bamberger rearrangement but suggests that PnbB first acts to effect a Bamberger-like rearrangement of 4HAB to 4A3HB followed by the replacement of the 4-amino group by a hydroxyl to form protocatechuate and that the phenoxazinone is produced as a result of some misrouting of the intermediate 4A3HB from its active site.

Ammonia-Lyases↗

PubMed2002

A third transposable element, ISPpu12, from the toluene-xylene catabolic plasmid pWW0 of Pseudomonas putida mt-2.

A 3,372-bp insertion sequence, ISPpu12, has been identified on the archetypal toluene-xylene TOL catabolic plasmid pWW0 from Pseudomonas putida mt-2. The insertion sequence element is located on the plasmid between bases 84397 and 87768 in a region which also contains the termini and transposase genes of the catabolic transposons Tn4651 and Tn4653 (A. Greated, L. Lambertson, P. A. Williams, and C. M. Thomas, Environ. Microbiol., in press). ISPpu12 has terminal inverted repeats of 24 bp with three mismatches and contains four open reading frames, a tnpA homologue and three open reading frames (lspA, orf1, and orf2) of undetermined function. After insertion in vitro of a Km(r) cassette into ISPpu12 either in the intergenic region between orf1 and orf2 or directly into the orf1 gene and ligation into a suicide vector, the modified ISPpu12-Km transposes at high frequency, often in multiple copies, into the chromosome of a P. putida recipient. Inactivation of lspA, orf1, and orf2 by introducing a 7-bp deletion into the 5' region of each gene had no major effect upon transposition, but a similar mutation of tnpA completely eliminated transposition. Analysis of the literature and of strains derived from the chlorobenzoate-degrading Pseudomonas sp. strain B13 suggests that the promiscuity of this element has played an important role in the history of plasmid pWW0. Database comparisons and the accompanying paper (A. J. Weightman, A. W. Topping, K. E. Hill, L. L. Lee, K. Sakai, J. H. Slater, and A. W. Thomas, J. Bacteriol. 184:6581-6591, 2002) show that ISPpu12 is a transposable element also found in other bacteria.

Bacterial Proteins↗

PubMed2002

Salicylate 5-hydroxylase from Ralstonia sp. strain U2: a monooxygenase with close relationships to and shared electron transport proteins with naphthalene dioxygenase.

The genes from the oxygenase cluster nagAaGHAbAcAd of naphthalene-degrading Ralstonia sp. strain U2 were cloned and overexpressed. Salicylate 5-hydroxylase (S5H) activity, converting salicylate to gentisate, was present in vitro only in the single extract of cells with overexpressed nagAaGHAb or in a mixture of three cell extracts containing, respectively, NagGH (the oxygenase components), NagAa (ferredoxin reductase), and NagAb (ferredoxin). Each of the three extracts required for S5H activity was rate limiting in the presence of excess of the others but, when in excess, did not affect the rate of catalysis. S5H catalyzed the 5-hydroxylation of the aromatic rings of 3- and 4-substituted salicylates. However, the methyl group of 5-methylsalicylate was hydroxylated to produce the 5-hydroxymethyl derivative and the 6-position on the ring of 5-chlorosalicylate was hydroxylated, producing 5-chloro-2,6-dihydroxybenzoate. In an assay for the nag naphthalene dioxygenase (NDO) based on the indole-linked oxidation of NADH, three extracts were essential for activity (NagAcAd, NagAa, and NagAb). NDO and S5H were assayed in the presence of all possible combinations of the nag proteins and the corresponding nah NDO proteins from the "classical" naphthalene degrader P. putida NCIMB9816. All three oxygenase components functioned with mixed combinations of the electron transport proteins from either strain. The S5H from strain U2 is a unique monooxygenase which shares sequence similarity with dioxygenases such as NDO but is also sufficiently similar in structure to interact with the same electron transport chain and probably does so in vivo during naphthalene catabolism in strain U2.

Dioxygenases↗

PubMed1998

Implications of the xylQ gene of TOL plasmid pWW102 for the evolution of aromatic catabolic pathways.

Pseudomonas putida strain O2C2 is able to grow on toluene, m-xylene and p-xylene through benzoate and the corresponding methylbenzoates (toluates). The catabolic genes are encoded on a large TOL plasmid, pWW102, of > 220 kb. The complete catabolic genes were cloned on four large overlapping restriction fragments covering a total of 28 kb of the plasmid, which was carefully mapped by restriction enzyme analysis. The presence of the xyl genes on the cloned DNA was confirmed by assay of representative enzymes of both operons. Virtually all the genes were located on the cloned DNA by hybridization of Southern blots with gene-specific probes from related pathways of other catabolic plasmids. Within the limitations of available restriction sites, the analysis showed that the genes are in two blocks. The major block carries the meta pathway operon xylXYZLTEGFJQKIH with the two regulatory genes xylSR immediately downstream. The upper pathway operon xylUWCMAB(N) is about 2-3 kb downstream of the regulatory genes and transcribed in the same direction as the meta pathway operon. Within each operon the gene order appears to be identical to that found in other TOL plasmids, but the relative location of the operons most closely resembles that found on plasmid pWW53, although there is no evidence of any xyl duplications on pWW102. The nucleotide sequence of the xylQ gene for the acetaldehyde dehydrogenase (acylating; ADA), together with the 3'-end of the upstream xylJ (for 2-oxopent-4-enoate hydratase) and the 5'-end of the downstream xylK (for 4-hydroxy-2-oxovalerate aldolase), was determined. The xylQ gene was ligated into expression vector pTrc99a and high levels of XylQ protein were detected by enzyme assay and by SDS-PAGE. All three genes xylJQK showed a high degree of homology with genes encoding isofunctional proteins from other Pseudomonas meta pathways, the highest being with the naphthalene catabolic genes nahLOM from the plasmid of Pseudomonas sp. NCIB 9816. The implications of the sequence homologies to the evolution of these pathways are discussed.

Aldehyde Oxidoreductases↗