bluuebunny/crossref_metadata_2025_split huggingface dataset:
Prediction of aggregation rate and aggregation‐prone segments in polypeptide sequences
Authors: Tartaglia Gian Gaetano , Cavalli Andrea , Pellarin Riccardo , Caflisch Amedeo 
Published: September 2005

AbstractThe reliable identification of β‐aggregating stretches in protein sequences is essential for the development of therapeutic agents for Alzheimer's and Parkinson's diseases, as well as other pathological conditions associated with protein deposition. Here, a model based on physicochemical properties and computational design of β‐aggregating peptide sequences is shown to be able to predict the aggregation rate over a large set of natural polypeptide sequences. Furthermore, the model identifies aggregation‐prone fragments within proteins and predicts the parallel or anti‐parallel β‐sheet organization in fibrils. The model recognizes different β‐aggregating segments in mammalian and nonmammalian prion proteins, providing insights into the species barrier for the transmission of the prion disease.
A mutation designed to alter crystal packing permits structural analysis of a tight‐binding fluorescein–scFv complex
Authors: Honegger Annemarie , Spinelli Silvia , Cambillau Christian , Plückthun Andreas 
Published: September 2005

AbstractThe structure of the scFv fragment FITC‐E2, obtained from a naive phage antibody scFv library derived from human donors, was determined at 2.1 Å resolution in the free form and at 3.0 Å in the complexed form. The wild‐type (wt) scFv binds fluorescein with a KD of 0.75 nM. The free scFv readily crystallizes by compacting its 18 amino acid‐long CDR‐H3, partially occluding the binding site and further blocking access by binding to the ”bottom” of a neighboring scFv molecule with a cluster of exposed aromatic residues within CDR‐H3. Only upon mutating one of the residues involved in this dominant crystal contact, an exposed tryptophan in the middle of CDR‐H3, crystals of the complex could be obtained. A series of alanine mutants within the putative antigen binding site, covering a range of binding affinities, were used to relate macroscopic thermodynamic and kinetic binding parameters to single‐molecule disruption forces measured by AFM. The effects of the mutations on the binding properties, particularly on the fraction of binding‐competent molecules within the population, cannot be fully explained by changes in the strength of local interactions. The significant conformational change of CDR‐H3 between the free and the liganded form illustrates the plasticity of the binding site. An accompanying study in this issue by Curcio and colleagues presents the molecular dynamics simulation of the forced unbinding experiments and explores possible effects of the mutations on the unbinding pathway of the hapten.
Two crystal forms of the restriction enzyme MspI–DNA complex show the same novel structure
Authors: Xu Qian Steven , Roberts Richard J. , Guo Hwai‐Chen 
Published: September 2005

AbstractThe crystal structure of the Type IIP restriction endonuclease MspI bound to DNA containing its cognate recognition sequence has been determined in both monoclinic and orthorhombic space groups. Significantly, these two independent crystal forms present an identical structure of a novel monomer–DNA complex, suggesting a functional role for this novel enzyme–DNA complex. In both crystals, MspI interacts with the CCGG DNA recognition sequence as a monomer, using an asymmetric mode of recognition by two different structural motifs in a single polypeptide. In the crystallographic asymmetric unit, the two DNA molecules in the two MspI–DNA complexes appear to stack with each other forming an end‐to‐end pseudo‐continuous 19‐mer duplex. They are primarily B‐form and no major bends or kinks are observed. For DNA recognition, most of the specific contacts between the enzyme and the DNA are preserved in the orthorhombic structure compared with the monoclinic structure. A cation is observed near the catalytic center in the monoclinic structure at a position homologous to the 74/45 metal site of EcoRV, and the orthorhombic structure also shows signs of this same cation. However, the coordination ligands of the metal are somewhat different from those of the 74/45 metal site of EcoRV. Combined with structural information from other solved structures of Type II restriction enzymes, the possible relationship between the structures of the enzymes and their cleavage behaviors is discussed.
Crystal structures of oxidized and reduced forms of human mitochondrial thioredoxin 2
Authors: Smeets Aude , Evrard Christine , Landtmeters Marie , Marchand Cécile , Knoops Bernard , Declercq Jean‐Paul 
Published: September 2005

AbstractMammalian thioredoxin 2 is a mitochondrial isoform of highly evolutionary conserved thioredoxins. Thioredoxins are small ubiquitous protein–disulfide oxidoreductases implicated in a large variety of biological functions. In mammals, thioredoxin 2 is encoded by a nuclear gene and is targeted to mitochondria by a N‐terminal mitochondrial presequence. Recently, mitochondrial thioredoxin 2 was shown to interact with components of the mitochondrial respiratory chain and to play a role in the control of mitochondrial membrane potential, regulating mitochondrial apoptosis signaling pathway. Here we report the first crystal structures of a mammalian mitochondrial thioredoxin 2. Crystal forms of reduced and oxidized human thioredoxin 2 are described at 2.0 and 1.8 Å resolution. Though the folding is rather similar to that of human cytosolic/nuclear thioredoxin 1, important differences are observed during the transition between the oxidized and the reduced states of human thioredoxin 2, compared with human thioredoxin 1. In spite of the absence of the Cys residue implicated in dimer formation in human thioredoxin 1, dimerization still occurs in the crystal structure of human thioredoxin 2, mainly mediated by hydrophobic contacts, and the dimers are associated to form two‐dimensional polymers. Interestingly, the structure of human thioredoxin 2 reveals possible interaction domains with human peroxiredoxin 5, a substrate protein of human thioredoxin 2 in mitochondria.
The leukocidin pore: Evidence for an octamer with four LukF subunits and four LukS subunits alternating around a central axis
Authors: Jayasinghe Lakmal , Bayley Hagan 
Published: September 2005

AbstractThe staphylococcal α‐hemolysin (αHL) and leukocidin (Luk) polypeptides are members of a family of related β‐barrel pore‐forming toxins. Upon binding to susceptible cells, αHL forms water‐filled homoheptameric transmembrane pores. By contrast, Luk pores are formed by two classes of subunit, F and S, rendering a heptameric structure displeasing on symmetry grounds at least. Both the subunit stoichiometry and arrangement within the Luk pore have been contentious issues. Here we use chemical and genetic approaches to show that (1) the predominant, or perhaps the only, form of the Luk pore is an octamer; (2) the subunit stoichiometry is 1:1; and (3) the subunits are arranged in an alternating fashion about a central axis of symmetry, at least when a fused LukS‐LukF construct is used. The experimental approaches we have used also open up new avenues for engineering the arrangement of the subunits of β‐barrel pore‐forming toxins.
Delineation of xenobiotic substrate sites in rat glutathione S‐transferase M1‐1
Authors: Hearne Jennifer L. , Colman Roberta F. 
Published: September 2005

AbstractGlutathione S‐transferases catalyze the conjugation of glutathione with endogenous and exogenous xenobiotics. Hu and Colman (1995) proposed that there are two distinct substrate sites in rat GST M1‐1, a 1‐chloro‐2,4‐dintrobenzene (CDNB) substrate site located in the vicinity of tyrosine‐115, and a monobromobimane (mBBr) substrate site. To determine whether the mBBr substrate site is distinguishable from the CDNB substrate site, we tested S‐(hydroxyethyl)bimane, a nonreactive derivative of mBBr, for its ability to compete kinetically with the substrates. We find that S‐(hydroxyethyl)bimane is a competitive inhibitor (KI = 0.36 μM) when mBBr is used as substrate, but not when CDNB is used as substrate, demonstrating that these two sites are distinct. Using site‐directed mutagenesis, we have localized the mBBr substrate site to an area midway through α‐helix 4 (residues 90–114) and have identified residues that are important in the enzymatic reaction. Substitution of alanine at positions along α‐helix 4 reveals that mutations at positions 103, 104, and 109 exhibit a greater perturbation of the enzymatic reaction with mBBr than with CDNB as substrate. Various other substitutions at positions 103 and 104 reveal that a hydrophobic residue is necessary at each of these positions to maintain optimal affinity of the enzyme for mBBr and preserve the secondary structure of the enzyme. Substitutions at position 109 indicate that this residue is important in the enzyme's affinity for mBBr but has a minimal effect on Vmax. These results demonstrate that the promiscuity of rat GST M1‐1 is in part due to at least two distinct substrate sites.
Effects of cytochrome P450 3A modulators ketoconazole and carbamazepine on quetiapine pharmacokinetics
Authors: Grimm Scott W. , Richtand Neil M. , Winter Helen R. , Stams Karen R. , Reele Stots B. 
Published: September 2005

AimsTo explore the potential for drug interactions on quetiapine pharmacokinetics using in vitro and in vivo assessments.MethodsThe CYP enzymes responsible for quetiapine metabolite formation were assessed using recombinant expressed CYPs and CYP‐selective inhibitors. P‐glycoprotein (Pgp) transport was tested in MDCK cells expressing the human MDR1 gene. The effects of CYP3A4 inhibition were evaluated clinically in 12 healthy volunteers that received 25 mg quetiapine before and after 4 days of treatment with ketoconazole 200 mg daily. To assess CYP3A4 induction in vivo, 18 patients with psychiatric disorders were titrated to steady‐state quetiapine levels (300 mg twice daily), then titrated to 600 mg daily carbamazepine for 2 weeks.ResultsCYP3A4 was found to be responsible for formation of quetiapine sulfoxide and N‐ and O‐desalkylquetiapine and not a Pgp substrate. In the clinical studies, ketoconazole increased mean quetiapine plasma Cmax by 3.35‐fold, from 45 to 150 ng ml−1 (mean Cmax ratio 90% CI 2.51, 4.47) and decreased its clearance (Cl/F) by 84%, from 138 to 22 l h−1 (mean ratio 90% CI 0.13, 0.20). Carbamazepine decreased quetiapine plasma Cmax by 80%, from 1042 to 205 ng ml−1 (mean Cmax ratio 90% CI 0.14, 0.28) and increased its clearance 7.5‐fold, from 65 to 483 l h−1 (mean ratio 90% CI 6.04, 9.28).ConclusionsCytochrome P450 3A4 is a primary enzyme responsible for the metabolic clearance of quetiapine. Quetiapine pharmacokinetics were affected by concomitant administration of ketoconazole and carbamazepine, and therefore other drugs and ingested natural products that strongly modulate the activity or expression of CYP3A4 would be predicted to change exposure to quetiapine.
Strikingly higher interleukin (IL)-1α, IL-1β and soluble interleukin-1 receptor antagonist (sIL-1RA) but similar IL-2, sIL-2R, IL-3, IL-4, IL-6, sIL-6R, IL-10, tumour necrosis factor (TNF)-α, transforming growth factor (TGF)-β2 and interferon IFN-γ urine levels in healthy females compared to healthy males: protection against urinary tract injury?
Authors: Sadeghi M , Daniel V , Naujokat C , Weimer R , Opelz G 
Published: September 2005

SummaryThe aim of this prospective study was to examine gender-related differences of cytokines in the plasma and urine of healthy individuals that might provide a clue concerning the lower rate of chronic renal diseases in females. Soluble interleukin-1 receptor antagonist (sIL-1RA), interleukin (IL)-1α, IL-1β, IL-2, sIL-2R, IL-3, IL-4, IL-6, sIL-6R, IL-10, tumor necrosis factor (TNF)-α, transforming growth factor (TGF)-β2 and interferon (IFN)-γ were determined using standard enzyme-linked immunosorbent assay (ELISA). Cytokine levels were determined in simultaneously obtained plasma and urine samples of 18 male and 28 female healthy members of our laboratory staff. Urine cytokine levels were studied three times at 1-month intervals. All individuals had a negative urine nitrite test and showed no symptoms of urinary tract infection (UTI). Plasma levels of all studied cytokines were similar in males and females (P = n.s.). However, females had significantly higher urine IL-1α (P &lt; 0·0001; P &lt; 0·0001; P &lt; 0·0001) and sIL-1RA (P = 0·0001; P = 0·0003; P = 0·0002) than males at three and higher IL-1β at one of the three investigations (P = 0·098; P = 0·003; P = 0·073). Urine levels of the other cytokines were similar in males and females. Higher urine levels of IL-1α, IL-1β and sIL-1RA in females may result from stimulation of cells in the urinary tract. Increased sIL-1RA might block T lymphocyte activation. The elevated cytokines may play a role in the protection of the female urinary tract from certain renal diseases, such as pyelonephritis and other inflammatory and sclerotic kidney diseases.
Cytokine responses correlate differentially with age in infancy and early childhood
Authors: Härtel C , Adam N , Strunk T , Temming P , Müller-Steinhardt M , Schultz C 
Published: September 2005

Summary
The functional differentiation of immune cells at early age plays a central role in immune physiology, e.g. for the sufficient eradication of pathogens. However, imbalances in effector cell responses may also have an impact in the pathophysiology of childhood diseases such as atopy and autoimmune disorders. As information on immune cell responses in infancy and early childhood is scarce, we conducted an observational, cross-sectional study in healthy newborns (n = 18), infants and young children (n = 54) aged 1–96 months and adult controls (n = 19) to assess cytokine mRNA and protein expression upon phorbol 12-myristate 13-actate/ionomycin stimulation and LPS-induced IL-12 expression in monocytes. The intracellular expression of interferon (IFN)-γ, tumour necrosis factor (TNF)-α (R = 0·748, P &lt; 0·0001; R = 0·784, P &lt; 0·0001, respectively) and interleukin (IL)-2 protein expression (R = 0·384, P = 0·008) was demonstrated to increase progressively with age. While a correlation between IL-4 protein expression and age was noted (R = 0·342, P = 0·007), the levels of IL-5 and IL-10 protein expression tended to be regulated on an individual basis during infancy and early childhood. An age correlation was also observed for intracellular IL-12 expression (R = 0·331, P = 0·009) in monocytes. These findings are valuable for further assessment of normal variations and maturation processes in immune cell responses and for the clinical–therapeutic monitoring of immunological status in various childhood diseases.
Interferon-α2a is sufficient for promoting dendritic cell immunogenicity
Authors: Tamir A , Jordan W J , Ritter M , Habib N , Lechler R I , Foster G R , Lombardi G 
Published: September 2005

SummaryType I interferons (IFNs) are widely used therapeutically. IFN-α2a in particular is used as an antiviral agent, but its immunomodulatory properties are poorly understood. Dendritic cells (DCs) are the only antigen-presenting cells able to prime naive T cells and therefore play a crucial role in initiating the adaptive phase of the immune response. We studied the effects of IFN-α2a on DC maturation and its role in determining Th1/Th2 equilibrium. We found that IFN-α2a induced phenotypic maturation of DCs and increased their allostimulatory capacity. When dendritic cells were stimulated simultaneously by CD40 ligation and IFN-α2a, the production of interleukin (IL)-10 and IL-12 was increased. In contrast, lipopolysaccharide (LPS) stimulation in the presence of IFN-α2a mainly induced IL-10 release. The production of IFN-γ and IL-5 by the responder naive T cells was also amplified in response to IFN-α2a-treated DCs. Furthermore, IL-12 production by IFN-α2a-treated DCs was enhanced further in the presence of anti-IL-10 antibody. Different results were obtained when DCs were treated simultaneously with IFN-α2a and other maturation factors, in particular LPS, and then stimulated by CD40 ligation 36 h later. Under these circumstances, IFN-α2a did not modify the DC phenotype, and the production of IL-10/IL-12 and IFN-γ/IL-5 by DCs and by DC-stimulated naive T cells, respectively, was inhibited compared to the effects on DCs treated with maturation factors alone. Altogether, this work suggests that IFN-α2a in isolation is sufficient to promote DC activation, however, other concomitant events, such as exposure to LPS during a bacterial infection, can inhibit its effects. These results clarify some of the in vivo findings obtained with IFN-α2a and have direct implications for the design of IFN-α-based vaccines for immunotherapy.
Screening of anEchinococcus granulosuscDNA library with IgG4 from patients with cystic echinococcosis identifies a new tegumental protein involved in the immune escape
Authors: Ortona E , Margutti P , Delunardo F , Nobili V , Profumo E , Riganò R , Buttari B , Carulli G , Azzarà A , Teggi A , Bruschi F , Siracusano A 
Published: September 2005

SummaryThe worldwide problem of chronic Echinococcus granulosus disease calls for new parasite-derived immunomodulatory molecules. By screening an E. granulosus cDNA library with IgG4 from patients with active cystic echinococcosis, we identified a cDNA that encodes a predicted partial protein that immunofluorescence studies localized in the protoscolex tegument and on the germinal layer of cyst wall. We named this protein EgTeg because the 105 amino acid sequence scored highest against a family of Schistosoma tegumental proteins. Evaluating the role of EgTeg in the human early inflammatory response we found that EgTeg significantly inhibited polymorphonuclear cell (PMN) chemotaxis. Cytometric analysis of intracellular cytokines disclosed a significantly higher percentage of cells producing IL-4 than IFN-γ (P = 0·001, Student's t-test) in T lymphocytes from patients with cystic echinococcosis stimulated with EgTeg. EgTeg induced weak Th1-dependent proliferation in 42% of patients’ peripheral blood mononuclear cells. In immunoblotting (IB) analysis of total IgG and IgG subclass responses to EgTeg in patients with cystic echinococcosis, patients with other parasitoses, patients with cystic lesions and healthy controls, total IgG specific to EgTeg yielded high sensitivity (73%) but low specificity (44%) precluding its use in immunodiagnosis. Conversely, IgG4 specific to EgTeg gave acceptable sensitivity (65%) and high specificity (89%) suggesting its use in immunodiagnosis to confirm ultrasound documented cysts suggestive of E. granulosus. Because the new tegumental antigen EgTeg inhibits chemotaxis, induces IL-4-positive T lymphocytes and noncomplement fixing antibodies (IgG4) it is an immunomodulatory molecule associated with chronic infection.
Grazer species effects on epilithon nutrient composition
Authors: EVANS‐WHITE MICHELLE A. , LAMBERTI GARY A. 
Published: October 2005

Summary1. Field and laboratory experiments were conducted to investigate the excretion stoichiometry of nitrogen (N) and phosphorus (P) of two benthic macroinvertebrate grazers, the crayfish Orconectes propinquus and the snail Elimia livescens, that differ in body stoichiometry (mean body molar N : P 18 and 28, respectively). Crayfish excretion had a significantly higher ammonium : soluble reactive phosphorus (SRP) ratio in the laboratory and in three natural streams than did snails, as predicted by ecological stoichiometry theory.2. In greenhouse recirculating artificial streams, treatments consisting of crayfish, snails, or no grazers were used to examine responses in dissolved nutrient concentrations and epilithon nutrient composition and limitation. SRP concentrations depended upon the grazer species, with the snail treatment having a higher SRP concentration than other treatments (P < 0.05). Dissolved inorganic N was not affected by grazers, but appeared to be rapidly incorporated in epilithon.3. Epilithon N content was dependent upon the grazer species present, with the crayfish treatment having a significantly higher N content than other treatments (P = 0.001). No grazer species effects on epilithon P content were found. However, both grazer treatments had significantly lower epilithon P content than the no‐grazer treatment.4. Traditionally, studies have focused on how grazer‐induced structural changes to epilithon can alter epilithon nutrient dynamics, but this structural mechanism could not solely explain differences in epilithon nutrient contents and ratios in the present study. Our results rather suggest that benthic grazers can alter epilithon nutrient composition and limitation via nutrient excretion. Consequently, macroinvertebrate grazers may serve as ‘nutrient pumps’ that partly regulate the availability of nutrients to algae in stream ecosystems.
Human exposure assessment to environmental chemicals using biomonitoring
Authors: CALAFAT ANTONIA M. , YE XIAOYUN , SILVA MANORI J. , KUKLENYIK ZSUZSANNA , NEEDHAM LARRY L. 
Published: September 2005

SummaryIn modern societies, humans may be exposed to a wide spectrum of environmental chemicals. Although the health significance of this exposure for many chemicals is unknown, studies to investigate the prevalence of exposure are warranted because of the chemicals’ potential harmful health effects, as often indicated in animal studies. Three tools have been used to assess exposure: exposure history/questionnaire information, environmental monitoring, and biomonitoring (i.e. measuring concentrations of the chemicals, their metabolites, or their adducts in human specimens). We present an overview on the use of biomonitoring in exposure assessment using phthalates, bisphenol A and other environmental phenols, and perfluorinated chemicals as examples. We discuss some factors relevant for interpreting and understanding biomonitoring data, including selection of both biomarkers of exposure and human matrices, and toxicokinetic information. The use of biomonitoring in human risk assessment is not discussed.