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D C Rees

Publications and source records attributed to D C Rees.

At least 91 records · Page 5Linked to original sources
PubMed1996

Structure of the R65Q mutant of yeast 3-phosphoglycerate kinase complexed with Mg-AMP-PNP and 3-phospho-D-glycerate.

The structure of a ternary complex of the R65Q mutant of yeast 3-phosphoglycerate kinase (PGK) with magnesium 5'-adenylylimidodiphosphate (Mg-AMP-PNP) and 3-phospho-D-glycerate (3-PG) has been determined by X-ray crystallography to 2.4 angstrom resolution. The structure was solved by single isomorphous replacement, anamalous scattering, and solvent flattening and has been refined to an R-factor of 0.185, with rms deviations from ideal bond distance and angles of 0.009 angstrom and 1.78 degrees, respectively. PGK consists of two domains, with the 3-PG bound to a "basic patch" of residues from the N-terminal domain and the Mg-AMP-PNP interacting with residues from the C-terminal domain. The two ligands are separated by approximately 11 angstrom across the interdomain cleft. The model of the R65Q mutant of yeast PGK is very similar to the structures of PGK isolated from horse, pig, and Bacillus stearothermophilus (rms deviations between equivalent alpha-carbons in the individual domains < 1.0 angstrom) but exhibits substantial variations with a previously reported yeast structure (rms deviations between equivalent alpha-carbons in the individual domains of 2.9-3.2 angstrom). The most significant tertiary structural differences among the yeast R65Q, equine, porcine, and B. stearothermophilus PGK structures occur in the relative orientations of the two domains. However, the relationships between the observed conformations of PGK are inconsistent with a "hinge-bending" behavior that would close the interdomain cleft. It is proposed that the available structural and biochemical data on PGK may indicate that the basic patch primarily represents the site of anion activation and not the catalytically active binding site for 3-PG.

Adenylyl Imidodiphosphate↗
PubMed1996

Heparin structure and interactions with basic fibroblast growth factor.

Crystal structures of heparin-derived tetra- and hexasaccharides complexed with basic fibroblast growth factor (bFGF) were determined at resolutions of 1.9 and 2.2 angstroms, respectively. The heparin structure may be approximated as a helical polymer with a disaccharide rotation of 174 degrees and a translation of 8.6 angstroms along the helix axis. Both molecules bound similarly to a region of the bFGF surface containing residues asparagine-28, arginine-121, lysine-126, and glutamine-135, the hexasaccharide also interacted with an additional binding site formed by lysine-27, asparagine-102, and lysine-136. No significant conformational change in bFGF occurred upon heparin oligosaccharide binding, which suggests that heparin primarily serves to juxtapose components of the FGF signal transduction pathway.

Binding Sites↗
PubMed1996

Evidence for a single origin of factor V Leiden.

Factor V Leiden is the most common hereditary blood clotting disorder so far identified, with an allele frequency of 4%. The low prevalence of the mutation outside of Europe suggests it occurred as a single event in the European founding population. In this study four polymorphisms have been identified defining different haplotypes in the exon 13 region of the factor V gene. One of these polymorphisms predicts a novel amino acid change, threonine to serine, in the B-domain of factor V. Statistical evidence for a single origin of factor V Leiden is provided through the association of the mutation with a single exon 13 haplotype.

Base Sequence↗
PubMed1996

Hb Sun Prairie: diagnostic pitfalls in thalassemic hemoglobinopathies.

We report an Asian Indian family in which two daughters have Hb Sun Prairie, a known unstable alpha 2-globin variant [codon 130, GCT-->CCT; alpha 2 130(H13)Ala-->Pro beta 2]. While the homozygous probands have chronic hemolysis-the same phenotype as previously reported, the heterozygous parents are asymptomatic with a thalassemia carrier phenotype, distinct from the chronic hemolytic state previously described in a heterozygote. Unlike the earlier cases in which family studies were not available, this family clearly exhibits autosomal recessive inheritance, unusual amongst variants within the same region of helix H. Globin chain biosynthesis ratios initially suggested a beta-thalassemic hemoglobinopathy-this was excluded by normal sequence analysis of both beta-globin genes. This case report further illustrates the complexity of phenotypes in the thalassemic hemoglobinopathies. It also demonstrates inversion of the alpha/beta-globin chain biosynthesis ratio, a phenomenon which had been noted in other alpha-globin variants and can be a confounding factor in the investigation of thalassemic hemoglobinopathies.

Adolescent↗
PubMed1995

World distribution of factor V Leiden.

We have analysed 3380 chromosomes (1690 unrelated individuals) from twenty-four populations for the presence of factor V Leiden, an important risk factor in venous thromboembolism. The allele frequency in 618 Europeans was 4.4%, with the highest prevalence among Greeks (7%). It was 0.6% in Asia Minor. Factor V Leiden was not found in any of 1600 chromosomes from Africa, Southeast Asia, Australasia, and the Americas. This distribution may partly explain the rarity of thromboembolic disease in these populations. The high prevalence in Europeans suggests that screening for this mutation should be considered in some circumstances.

Africa↗
PubMed1995

Crystal structure of a conserved protease that binds DNA: the bleomycin hydrolase, Gal6.

Bleomycin hydrolase is a cysteine protease that hydrolyzes the anticancer drug bleomycin. The homolog in yeast, Gal6, has recently been identified and found to bind DNA and to act as a repressor in the Gal4 regulatory system. The crystal structure of Gal6 at 2.2 A resolution reveals a hexameric structure with a prominent central channel. The papain-like active sites are situated within the central channel, in a manner resembling the organization of active sites in the proteasome. The Gal6 channel is lined with 60 lysine residues from the six subunits, suggesting a role in DNA binding. The carboxyl-terminal arm of Gal6 extends into the active site cleft and may serve a regulatory function. Rather than each residing in distinct, separable domains, the protease and DNA-binding activities appear structurally intertwined in the hexamer, implying a coupling of these two activities.

Amino Acid Sequence↗
PubMed1995

Structure of a hyperthermophilic tungstopterin enzyme, aldehyde ferredoxin oxidoreductase.

The crystal structure of the tungsten-containing aldehyde ferredoxin oxidoreductase (AOR) from Pyrococcus furiosus, a hyperthermophilic archaeon (formerly archaebacterium) that grows optimally at 100 degrees C, has been determined at 2.3 angstrom resolution by means of multiple isomorphous replacement and multiple crystal form averaging. AOR consists of two identical subunits, each containing an Fe4S4 cluster and a molybdopterin-based tungsten cofactor that is analogous to the molybdenum cofactor found in a large class of oxotransferases. Whereas the general features of the tungsten coordination in this cofactor were consistent with a previously proposed structure, each AOR subunit unexpectedly contained two molybdopterin molecules that coordinate a tungsten by a total of four sulfur ligands, and the pterin system was modified by an intramolecular cyclization that generated a three-ringed structure. In comparison to other proteins, the hyperthermophilic enzyme AOR has a relatively small solvent-exposed surface area, and a relatively large number of both ion pairs and buried atoms. These properties may contribute to the extreme thermostability of this enzyme.

Aldehyde Oxidoreductases↗
PubMed1995

The importance of Arg40 and 45 in the mitogenic activity and structural stability of basic fibroblast growth factor: effects of acidic amino acid substitutions.

High-affinity binding of basic fibroblast growth factor (bFGF) to the tyrosine kinase receptor requires cell-surface heparan sulfate proteoglycan or exogenous addition of heparin. The crystal structure of bFGF shows Arg40 and 45 on the surface opposite to the heparin-binding region, suggesting that these charged residues may be involved in the receptor binding. Therefore, these amino acids were mutated to aspartic acid separately or simultaneously, and also a simultaneous mutation to glutamic acid was introduced. These mutants displayed a mitogenic activity decreased greater than tenfold compared to the wild-type protein. Addition of heparin had no effect on the activity, while these mutants showed heparin-binding characteristics resembling those of the native sequence protein. The mutants exhibited decreased stability compared to the native sequence protein. Gradual changes in conformation were observed by circular dichroic and infrared spectroscopy. Heparin chromatography also showed the presence of denatured form for these mutants. However, in the presence of multivalent anions such as citrate, sucrose octasulfate, and heparin, the conformation of the mutants resembled that of the wild-type protein, as revealed by X-ray crystallography and circular dichroism spectra of the mutant with a Arg40-->Asp substitution.

3T3 Cells↗
PubMed1995

A comparison of the mutagenicity of mainstream cigarette smoke condensates from a representative sample of the U.S. cigarette market with a Kentucky reference cigarette (K1R4F).

The Salmonella mutagenicity assay has been used to investigate the mutagenicity of cigarette smoke and cigarette smoke condensate. The Kentucky reference (K1R4F) cigarette is designed to be representative of full-flavor, low-tar cigarettes sold in the U.S. and to serve as a reference standard for comparative studies on the chemistry and biological activities of cigarette smoke and condensate. The objective of this study was to determine if the mutagenicity of mainstream smoke condensate from the K1R4F, as measured by the Salmonella mutagenicity assay, is representative of the mutagenic activity of U.S. cigarettes. Mainstream smoke condensates prepared in dimethyl sulfoxide from the K1R4F and 73 brand styles (representing greater than 70% of the total U.S. cigarette market) were assayed using Salmonella typhimurium TA98 and TA100 (+S9) at concentrations of 0, 25, 50, 75, 100, 125 and 250 micrograms/plate. Revertants/mg condensate were determined by calculating the slopes of the dose-response curves using linear and nonlinear regression models. Revertants/cigarette were determined by multiplying the revertants/mg condensate by the mg condensate/cigarette. No significant differences (p > 0.05) were observed between the mean mutagenicity of U.S. market and K1R4F mainstream smoke condensates in terms of revertants/mg condensate or revertants/cigarette. Increased variability in mutagenicity was observed among the U.S. brands versus that of the K1R4F. This is not surprising since variability among the U.S. brands would be expected to have both measurement error and brand style variability while the K1R4F variability contains only the measurement error portion. These results demonstrate that the K1R4F is a representative model for the U.S. cigarette market in comparative Salmonella mutagenicity studies using mainstream smoke condensates.

Mutagenicity Tests↗
PubMed1995

The metabolites of nitric oxide in sickle-cell disease.

Plasma NOx concentrations were raised in 22 acute painful crises in SCD. We have measured blood concentrations of nitric oxide metabolites (NOx) in sickle-cell disease (SCD), and shown that they are increased compared with healthy controls (P = 0.002), and haemoglobin E/beta-thalassaemic controls (P = 0.05). Concentrations in steady-state SCD were also higher than in healthy controls (P = 0.04) but not significantly different from the concentrations at the beginning of painful crises (P = 0.34). Importantly, in 12 regularly exchanged sicklers, the mean pre-transfusion NOx concentration did not differ significantly from the control population (P = 0.52), suggesting that the changes in NO metabolism can be reversed. It is unlikely that the increased concentrations of NOx in SCD result from anaemia or haemolysis as the untransfused haemoglobin E/beta-thalassaemics did not show increased levels.

Acute Disease↗
PubMed1995

Molecular characterization of the genes encoding the tungsten-containing aldehyde ferredoxin oxidoreductase from Pyrococcus furiosus and formaldehyde ferredoxin oxidoreductase from Thermococcus litoralis.

The hyperthermophilic archaea Pyrococcus furiosus and Thermococcus litoralis contain the tungstoenzymes aldehyde ferredoxin oxidoreductase, a homodimer, and formaldehyde ferredoxin oxidoreductase, a homotetramer. herein we report the cloning and sequencing of the P. furiosus gene aor (605 residues; M(r), 66,630) and the T. litoralis gene for (621 residues; M(r), 68,941).

Aldehyde Oxidoreductases↗
PubMed1995

Are serum concentrations of nitric oxide metabolites useful for predicting the clinical outcome of severe ulcerative colitis?

OBJECTIVE: To determine serum concentrations of nitric oxide metabolites (NOX) in patients with severe ulcerative colitis and to assess whether these concentrations predict clinical outcome. PATIENTS: Twenty-six patients (16 men and 10 women, mean age 46 years) with severe ulcerative colitis requiring hospitalization for parenteral steroid therapy. Thirteen patients had a complete clinical response and symptoms resolved after 5 days of parenteral steroid administration; 13 made an incomplete recovery and needed further treatment (six cyclosporin, seven colectomy). METHODS: Serum concentrations of NOX and C-reactive protein (CRP) were measured daily for 3 days in all patients and as clinically indicated thereafter. The normal range for NOX was established by measuring the concentration in 25 healthy controls. RESULTS: Mean serum NOX and CRP concentrations were significantly elevated in both the patients with a complete and those with an incomplete response compared with controls (P < 0.001) on day 1 and fell during the first 3 days of therapy. On day 3, mean serum concentrations of NOX and CRP were lower in the patients with a complete response, but only the difference in CRP attained statistical significance (P = 0.02). There was no correlation between NOX and CRP concentrations. CONCLUSIONS: In the majority of patients with severe ulcerative colitis, circulating concentrations of NOX are increased at presentation and fall promptly during parenteral steroid therapy, irrespective of clinical outcome. However, in a small number of patients NOX concentrations do not fall during steroid treatment and such patients will probably require additional medical therapy or surgery.

Adult↗