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C J Schofield

Publications and source records attributed to C J Schofield.

At least 55 records · Page 3Linked to original sources
PubMed2000

Oxidative degradation of bilirubin produces vasoactive compounds.

Subarachnoid haemorrhage is often followed by haemolysis and concomitant oxidative stress, and is frequently complicated by pathological vasoconstriction or cerebral vasospasm. It is known that upregulation of haem oxygenase (HO-1) is induced by oxidative stress and results in release of biliverdin and bilirubin (BR), which are scavengers of reactive oxygen species (ROS). Here we report biomimetic studies aimed at modelling pathological conditions leading to oxidative degradation of BR. Oxidative degradation products of BR, formed by reaction with hydrogen peroxide (an ROS model system), demonstrated biological activity by stimulating oxygen consumption and force development in vascular smooth muscle from porcine carotid artery. Analogous biological activity was observed with vasoactive cerebrospinal fluid from subarachnoid haemorrhage patients. Three degradation products of BR were isolated: two were assigned as isomeric monopyrrole (C9H11N2O2) derivatives, 4-methyl-5-oxo-3-vinyl-(1, 5-dihydropyrrol-2-ylidene)acetamide and 3-methyl-5-oxo-4-vinyl-(1, 5-dihydropyrrol-2-ylidene)acetamide and the third was 4-methyl-3-vinylmaleimide (MVM), a previously isolated photodegradation product of biliverdin. Possible mechanisms of oxidative degradation of BR are discussed. Tentative assignment of these structures in the cerebrospinal fluid (CSF) of cerebral vasospasm patients has been made. It is proposed that one or more of the degradation products of biliverdin or bilirubin are involved in complications such as vasospasm and or pathological vasoconstriction associated with haemorrhage.

Bilirubin↗
PubMed2000

Chromosome homogeneity in populations of Triatoma brasiliensis Neiva 1911 (Hemiptera - reduviidae - triatominae).

Triatoma brasiliensis is the most important vector of Chagas disease in the semiarid zone of the Northeast of Brazil. Several authors have reported the occurrence of four chromatic patterns with morphological, ecological, and genetic differences. In order to determine the existence of cytogenetic differentiation between these chromatic forms, we analyzed their karyotypes and the chromosome behavior during the male meiotic process. Triatoma brasiliensis shows distinct and specific chromosome characteristics, which differ from those observed in all other triatomine species. However, no cytogenetic differences were observed between the four chromatic forms of T. brasiliensis. The lack of chromosome differentiation among them could indicate that the populations of this species are in a process of differentiation that does not involve their chromosomal organization.

Animals↗
PubMed2000

Biosystematics and evolution of the Triatominae.

In this paper we summarize the systematics of the 130 currently recognized species of Triatominae and the key features of their evolutionary background. There is increasing evidence that the subfamily has polyphyletic origins, with the various tribes and species groups probably arising from different reduviid lineages in relatively recent times.

Adaptation, Physiological↗
PubMed2000

Genetic variability of Triatoma brasiliensis (Hemiptera: Reduviidae) populations.

Isoenzymes, random amplified polymorphic DNA (RAPD), and morphometry were used to compare genetic variability within and between three populations of Triatoma brasiliensis Neiva, 1911 from northeast Brazil. The isoenzyme profiles were identical for the three populations, whereas the complex RAPD patterns allowed clear discrimination between the three. Morphometric analysis, using characters of the head capsule, also showed discrimination between the three populations but only in comparisons of males, not females. The results show considerable genetic heterogeneity in T. brasiliensis, with an indication of geographic structuring possibly resulting from a recent series of domestication events.

Animals↗
PubMed2000

Phylogeny and molecular taxonomy of the Rhodniini derived from mitochondrial and nuclear DNA sequences.

Eleven species of Rhodnius and one of Psammolestes were compared by DNA sequence analysis of fragments of the mitochondrial large subunit ribosomal RNA (mtlsurRNA), the mitochondrial cytochrome b (mtCytb), and the D2 variable region of the 28S nuclear RNA (D2), totaling 1,429 base pairs. The inferred phylogeny, using Triatoma infestans as an outgroup, revealed two main clades within the Rhodniini--one, including the prolixus group of species (Rhodnius prolixus, Rhodnius robustus, Rhodnius neglectus, and Rhodnius nasutus) together with Rhodnius domesticus and Rhodnius neivai, and the other comprising two groups formed by Rhodnius pictipes plus Rhodnius brethesi, and Rhodnius ecuadoriensis plus Rhodnius pallescens. Psammolestes tertius appeared most closely related to the prolixus group. The analysis strongly supports the validity of R. robustus as a species distinct from others of the prolixus group, but suggests higher genetic structuring of R. robustus populations compared to the other species. Although R. robustus has been found naturally infected by Trypanosoma cruzi, the fact that it is apparently entirely sylvatic and unable to establish in homes suggests that it is of no great importance as a Chagas disease vector in humans.

Animals↗
PubMed2000

[Operational aspects of Triatoma brasiliensis control].

Vector control strategies against indigenous species is not easy, due to their capacity to reinvade treated premises from sylvatic ecotopes. Between August 1996 and December 1997 we conducted a study on reinfestation of houses after spraying in a county in the State of Ceará. Of 277 houses examined, 113 (40.8%) were infested (21.7% intradomiciliary and 35.4% peridomiciliary). Of the 433 Triatominae collected, 207 were Triatoma brasiliensis (49% of which intradomiciliary, with a mean of 1.8 insects/house) and 226 were Triatoma pseudomaculata (97% peridomiciliary). The age structure of the two indicated a univoltine development cycle for T. brasiliensis and two cycles per year for T. pseudomaculata. Four months after spraying with deltamethrin SC 25mg ia/m2, 9.7% of the houses were still positive, mainly with peridomestic infestations. Intradomiciliary wall bioassays showed persistence of the insecticide up to 9 months after spraying. Considering the high potential for recolonization of treated premises from sylvatic foci, we propose an operational strategy combining traditional evaluations and community-based surveillance with increased selective interventions and community education.

Animals↗
PubMed1999

Mechanistic insights into the inhibition of serine proteases by monocyclic lactams.

Although originally discovered as inhibitors of pencillin-binding proteins, beta-lactams have more recently found utility as serine protease inhibitors. Indeed through their ability to react irreversibly with nucleophilic serine residues they have proved extraordinarily successful as enzyme inhibitors. Consequently there has been much speculation as to the reason for the general effectiveness of beta-lactams as antibacterials or inhibitors of hydrolytic enzymes. The interaction of analogous beta- and gamma-lactams with a serine protease was investigated. Three series of gamma-lactams based upon monocyclic beta-lactam inhibitors of elastase [Firestone, R. A. et al. (1990) Tetrahedron 46, 2255-2262.] but with an extra methylene group inserted between three of the bonds in the ring were synthesized. Their interaction with porcine pancreatic elastase and their efficacy as inhibitors were evaluated through the use of kinetic, NMR, mass spectrometric, and X-ray crystallographic analyses. The first series, with the methylene group inserted between C-3 and C-4 of the beta-lactam template, were readily hydrolyzed but were inactive or very weakly active as inhibitors. The second series, with the methylene group between C-4 and the nitrogen of the beta-lactam template, were inhibitory and reacted reversibly with PPE to form acyl-enzyme complexes, which were stable with respect to hydrolysis. The third series, with the methylene group inserted between C-2 and C-3, were not hydrolyzed and were not inhibitors consistent with lack of binding to PPE. Comparison of the crystal structure of the acyl-enzyme complex formed between PPE and a second series gamma-lactam and that formed between PPE and a peptide [Wilmouth, R. C., et al. (1997) Nat. Struct. Biol. 4, 456-462.] reveals why the complexes formed with this series were resistant to hydrolysis and suggests ways in which stable acyl-enzyme complexes might be obtained from monocyclic gamma-lactam-based inhibitors.

Animals↗
PubMed1999

Studies on the active site of deacetoxycephalosporin C synthase.

The Fe(II) and 2-oxoglutarate-dependent dioxygenase deacetoxycephalosporin C synthase (DAOCS) from Streptomyces clavuligerus was expressed at ca 25 % of total soluble protein in Escherichia coli and purified by an efficient large-scale procedure. Purified protein catalysed the conversions of penicillins N and G to deacetoxycephems. Gel filtration and light scattering studies showed that in solution monomeric apo-DAOCS is in equilibrium with a trimeric form from which it crystallizes. DAOCS was crystallized +/-Fe(II) and/or 2-oxoglutarate using the hanging drop method. Crystals diffracted to beyond 1.3 A resolution and belonged to the R3 space group (unit cell dimensions: a=b=106.4 A, c=71.2 A; alpha=beta=90 degrees, gamma=120 degrees (in the hexagonal setting)). Despite the structure revealing that Met180 is located close to the reactive oxidizing centre of DAOCS, there was no functional difference between the wild-type and selenomethionine derivatives. X-ray absorption spectroscopic studies in solution generally supported the iron co-ordination chemistry defined by the crystal structures. The Fe K-edge positions of 7121.2 and 7121.4 eV for DAOCS alone and with 2-oxoglutarate were both consistent with the presence of Fe(II). For Fe(II) in DAOCS the best fit to the Extended X-ray Absorption Fine Structure (EXAFS) associated with the Fe K-edge was found with two His imidazolate groups at 1.96 A, three nitrogen or oxygen atoms at 2.11 A and one other light atom at 2.04 A. For the Fe(II) in the DAOCS-2-oxoglutarate complex the EXAFS spectrum was successfully interpreted by backscattering from two His residues (Fe-N at 1.99 A), a bidentate O,O-co-ordinated 2-oxoglutarate with Fe-O distances of 2.08 A, another O atom at 2.08 A and one at 2.03 A. Analysis of the X-ray crystal structural data suggests a binding mode for the penicillin N substrate and possible roles for the C terminus in stabilising the enzyme and ordering the reaction mechanism.

Binding Sites↗
PubMed1999

The Southern Cone Initiative against Chagas disease.

Chagas disease (also known as American trypanosomiasis) is now ranked as the most serious parasitic disease of the Americas, with an economic impact far outranking the combined effects of other parasitic diseases such as malaria, schistosomiasis and leishmaniasis. Although the chronic infection remains virtually incurable, transmission can be halted by eliminating the domestic insect vectors and screening blood donors to avoid transfusional transmission. In line with this strategy, governments of the six Southern Cone countries (Argentina, Bolivia, Brazil, Chile, Paraguay and Uruguay) launched in 1991 an ambitious initiative to control Chagas disease through elimination of the main vector, Triatoma infestans, and large-scale screening of blood donors. Now at its mid-point, the programme has achieved remarkable success, with transmission halted over vast areas of the previously endemic regions. Well over 2 million rural houses have been sprayed to eliminate T. infestans, and the programme has already shown significant economic rates of return in addition to the medical and social benefits.

Animals↗
PubMed1999

Structural and mechanistic studies on 2-oxoglutarate-dependent oxygenases and related enzymes.

Mononuclear nonheme-Fe(II)-dependent oxygenases comprise an extended family of oxidising enzymes, of which the 2-oxoglutarate-dependent oxygenases and related enzymes are the largest known subgroup. Recent crystallographic and mechanistic studies have helped to define the overall fold of the 2-oxoglutarate-dependent enzymes and have led to the identification of coordination chemistry closely related to that of other nonheme-Fe(II)-dependent oxygenases, suggesting related mechanisms for dioxygen activation that involve iron-mediated electron transfer.

Amino Acid Oxidoreductases↗
PubMed1999

Comparison of isoenzyme electrophoresis and morphometric analysis for phylogenetic reconstruction of the Rhodniini (Hemiptera: Reduviidae: Triatominae).

A phylogenetic reconstruction of the medically important tribe Rhodniini (Hemiptera: Reduviidae) based on multilocus isoenzyme electrophoresis is compared with phylogenetic patterns derived from a traditional morphometric analysis. Even with non-normality in the morphometric data, and some inequalities in population variances, discriminant analysis of size-free variables provided broadly similar phylogenetic information to that derived from isoenzyme analysis, revealing 3 main species groups within the genus Rhodnius.

Animals↗
PubMed1999

Triatominae as a model of morphological plasticity under ecological pressure.

The use of biochemical and genetic characters to explore species or population relationships has been applied to taxonomic questions since the 60s. In responding to the central question of the evolutionary history of Triatominae, i.e. their monophyletic or polyphyletic origin, two important questions arise (i) to what extent is the morphologically-based classification valid for assessing phylogenetic relationships? and (ii) what are the main mechanisms underlying speciation in Triatominae? Phenetic and genetic studies so far developed suggest that speciation in Triatominae may be a rapid process mainly driven by ecological factors.

Animals↗
PubMed1998

Inhibition of elastase by N-sulfonylaryl beta-lactams: anatomy of a stable acyl-enzyme complex.

beta-Lactam inhibitors of transpeptidase enzymes involved in cell wall biosynthesis remain among the most important therapeutic agents in clinical use. beta-Lactams have more recently been developed as inhibitors of serine proteases including elastase. All therapeutically useful beta-lactam inhibitors operate via mechanisms resulting in the formation of hydrolytically stable acyl-enzyme complexes. Presently, it is difficult to predict which beta-lactams will form stable acyl-enzyme complexes with serine enzymes. Further, the factors that result in the seemingly special nature of beta-lactams versus other acylating agents are unclear-if indeed they exist. Here we present the 1.6 A resolution crystal structure of a stable acyl-enzyme complex formed between porcine pancreatic elastase and a representative monocyclic beta-lactam, which forms a simple acyl-enzyme. The structure shows that the ester carbonyl is not located within the oxyanion hole and the "hydrolytic" water is displaced. Combined with additional kinetic and mass spectrometric data, the structure allows the rationalization of the low degree of hydrolytic lability observed for the beta-lactam-derived acyl-enzyme complex.

Amino Acid Sequence↗
PubMed1998

Structure of a cephalosporin synthase.

Penicillins and cephalosporins are among the most widely used therapeutic agents. These antibiotics are produced from fermentation-derived materials as their chemical synthesis is not commercially viable. Unconventional steps in their biosynthesis are catalysed by Fe(II)-dependent oxidases/oxygenases; isopenicillin N synthase (IPNS) creates in one step the bicyclic nucleus of penicillins, and deacetoxycephalosporin C synthase (DAOCS) catalyses the expansion of the penicillin nucleus into the nucleus of cephalosporins. Both enzymes use dioxygen-derived ferryl intermediates in catalysis but, in contrast to IPNS, the ferryl form of DAOCS is produced by the oxidative splitting of a co-substrate, 2-oxoglutarate (alpha-ketoglutarate). This route of controlled ferryl formation and reaction is common to many mononuclear ferrous enzymes, which participate in a broader range of reactions than their well-characterized counterparts, the haem enzymes. Here we report the first crystal structure of a 2-oxoacid-dependent oxygenase. High-resolution structures for apo-DAOCS, the enzyme complexed with Fe(II), and with Fe(II) and 2-oxoglutarate, were obtained from merohedrally twinned crystals. Using a model based on these structures, we propose a mechanism for ferryl formation.

Cloning, Molecular↗
PubMed1998

Alary polymorphism in Triatoma spinolai and its possible relationship with demographic strategy.

Among collections of Triatoma spinolai from various sites in northern Chile, adults from coastal populations are invariably wingless, whereas inland populations show balanced alary polymorphism between wingless females and males that are either winged or wingless. Laboratory crosses showed that male offspring from normal-winged parents were always winged (88% long-winged) and those from long-winged male parents were all long-winged. The male offspring from wingless males always included winged males: 11/33 = 33%, of which 8/11 = 73% were long-winged. An X-linked mutation is proposed to inhibit wing development. Field studies of population demography indicate that male alary polymorphism is advantageous in the desert environment of northern Chile.

Animals↗
PubMed1998

Population structure of Andean Triatoma infestans: allozyme frequencies and their epidemiological relevance.

Triatoma infestans (Hemiptera: Reduviidae) from 22 Andean localities in Bolivia (n = 968) and Peru (n = 37) were analysed by multi-locus enzyme electrophoresis. Among 12 gene-enzyme systems analysed, GPD, 6GPD and PGM were polymorphic, ACON, G6PD, GPI, 1DH, LAP, MDH, ME, PEP-A and PEP-B were monomorphic. Allozyme frequencies were analysed in relation to geographical and climatic factors, and the presence or absence of Trypanosoma cruzi infection. At one locality (Vallegrande, Bolivia), the frequency of 6Pgd-1 was significantly higher in infected (41% of 85) than in uninfected (17% of 83) adult T. infestans, although no such difference was found among nymphs (n = 347). From other localities, only insects infected with T. cruzi were subjected to isozyme analysis. Populations of T. infestans within villages showed panmixia, while genetic differentiation of T. infestans between villages was correlated with the distance between them. The genetic structure of T. infestans natural populations followed an 'isolation by distance' model, involving a series of founder effects followed by genetic drift, rather than adaptation in response to differential selection pressures. This conforms with circumstantial evidence that T. infestans spread, mainly in association with recent human migrations, from a source, probably in southern Bolivia. Isoenzyme characterization of populations of T. infestans could be used to infer sources of re-infestation during the surveillance phase of control programs.

Animals↗