bluuebunny/crossref_metadata_2025 huggingface dataset:
Prevalence of Celiac Disease in Brazilian Children with Type 1 Diabetes Mellitus
Authors: Tanure Mariella Guarino , Silva Ivani Novato , Bahia Magda , Penna Francisco José
Published: February 2006

Objective:

Although the relationship between celiac disease and diabetes mellitus type 1 is well recognized, there are no studies of this association in Brazil. This study aims to identify the prevalence of celiac disease in a group of children with diabetes mellitus type 1 undergoing treatment in the pediatric endocrinology division of a university hospital in Minas Gerais, Brazil.

Methods:

Immunoglobulin (Ig)A and IgG antigliadin antibodies (enzyme‐linked immunoadsorbent assay) were measured in blood collected from 236 children and adolescents with diabetes mellitus type 1. Patients with antigliadin antibodies then had jejunal biopsy and determination of antiendomysial antibodies by indirect immunofluorescence.

Results:

Twenty‐one patients had IgA or IgG antigliadin antibodies. Nineteen underwent jejunal biopsy. Six had mucosal alterations compatible with celiac disease; four had nonspecific histologic changes; nine had normal biopsies. Thirteen antigliadin antibody‐positive patients were antiendomysial antibody‐negative; one antiendomysial antibody‐negative patient had celiac disease. The prevalence of celiac disease was 2.6% among 234 patients.

Conclusions:

Measurement of antigliadin antibodies in patients with diabetes mellitus type 1 helped in the selection of patients to undergo jejunal biopsy. Antiendomysial antibodies were highly specific and moderately sensitive in predicting celiac disease. The prevalence of celiac disease was higher in diabetics than in the general population, suggesting the need for regular screening assessment of diabetic children.
Long‐Term Prognosis of Patients with Biliary Atresia: A 25 Year Summary
Authors: Hung Pei‐Yin , Chen Chiu‐Chiang , Chen Wei‐Jao , Lai Hong‐Shiee , Hsu Wen‐Ming , Lee Po‐Hung , Ho Ming‐Chih , Chen Tony H. H , Ni Yen‐Hsuan , Chen Huey‐Ling , Hsu Hong‐Yuan , Chang Mei‐Hwei
Published: February 2006

Objective:

The purpose of this study was to delineate the long‐term prognosis of biliary atresia (BA) in Taiwan.

Study Design:

From 1976 to 2000, 185 children were diagnosed with BA, 22 underwent exploratory laparotomy without Kasai operation, and 163 underwent Kasai operation, of which 141 cases had long‐term follow‐up and formed the basis of this study. The outcome was analyzed.

Results:

Among the 141 BA children studied who underwent Kasai operation, 115 (81.6%) had recoloration of stools, and 86 (61.0%) became jaundice‐free (bilirubin <20 μmol/L). The resolution of jaundice and the absence of repeated cholangitis contributed to better outcome. Five and 10 year survival rates with native liver were 35% and 31%, respectively. Liver transplantation was performed in 19 patients (all but 2 with a living‐related donor), and 15 (79%) survived. Five and 10 year overall survival rates for BA patients were 41.9% and 40.2%, respectively.

Conclusions:

The study delineated the long‐term outcome of BA in an Asian country other than Japan. Survival with native liver after a Kasai operation in Taiwan was similar to that in the American and European series. Limited donors for liver transplantation in the years of the study accounted for the poor overall prognosis of BA patients in this series.
Free Sugar and Sugar Alcohol Concentrations in Human Breast Milk
Authors: Cavalli Claudio , Teng Cecilia , Battaglia Frederick C , Bevilacqua Giulio
Published: February 2006

Objectives:

The goal was to determine the free sugars and polyols in human breast milk, both term and preterm, for comparison with formula milks.

Methods:

Methodology was developed for the measurement of the free sugars and polyols in breast milk. There were 16 samples collected from women who delivered at term and 17 samples from women delivering preterm. For purposes of comparison, samples were also collected from four commercial formulas for term infants and eight from commercial formulas for preterm infants as well as one sample of cows' milk. All samples were frozen immediately and analyzed by high‐performance liquid chromatography techniques.

Results:

Except for lactose concentrations, no significant differences were detected for all other sugars and polyols between term and preterm breast milk samples. Within breast milk samples, two patterns emerged, with one group containing additional elution peaks for compounds not yet identified. A second group did not contain these compounds. There were a number of significant differences between breast milk and formulas, particularly for inositol, glycerol, glucose, and galactose. All milks contained significant concentrations of mannose.

Conclusions:

There are no significant differences among breast milk samples for free sugar and polyol concentrations except lactose. However, some milk contains additional peaks that could be of dietary or genetic origin. Formula milk has relatively high concentrations of glucose and galactose compared with breast milk, suggesting some lactose hydrolysis.
Liver Abscesses in Children: A Single Center Experience in the Developed World
Authors: Muorah M , Hinds R , Verma A , Yu D , Samyn M , Mieli‐Vergani G , Hadžić N
Published: February 2006

Objectives:

The aim of this study was to investigate the clinical and radiologic features, predisposing risk factors, and complications of children with pyogenic liver abscess (PLA) referred to a tertiary pediatric hepatology center.

Methods:

We analyzed our database of all children referred to our unit over a 10 year period and performed a case note review of all patients with a radiologically proven PLA.

Results:

PLA was diagnosed in 15 children (7 boys), 0.5% of all referrals. They presented at a median age of 10 years (range 2 months‐15 years). In three children (2 boys), PLA was the first manifestation of chronic granulomatous disease. Among the others, five had radiologic evidence of other intra‐abdominal pathology (1 with subsequently proven appendicitis), and four developed portal vein thrombosis with portal hypertension. The commonest isolated pathogen was Staphylococcus aureus. Combined treatment with guided aspiration and prolonged intravenous antibiotics was successful in all patients.

Conclusion:

PLA is a rare diagnosis in children in the developed world. It may be caused by primary neutrophil disorders even in the absence of a previous history of infection. Co‐existent appendicitis, intra‐abdominal sepsis, and ascending pylephlebitis must be sought because these children are at risk of developing portal vein obstruction and portal hypertension. Prolonged intravenous antibiotic treatment guided by microbiologic sensitivities is highly effective.
Sodium Butyrate‐Mediated Sp3 Acetylation Represses Human Insulin‐Like Growth Factor Binding Protein‐3 Expression in Intestinal Epithelial Cells
Authors: White Nicholas R , Mulligan Peter , King Peter J , Sanderson Ian R
Published: February 2006

Objectives:

Butyrate concentrations in the gastrointestinal tract vary greatly with age. In intestinal epithelial cells, butyrate enhances gene transcription by increasing histone acetylation, rendering the nucleosome open to transcription factors. However, it inhibits human insulin‐like growth factor binding protein (hIGFBP)‐3 expression. We therefore hypothesized that butyrate also acts by regulating transcription factor acetylation.

Methods:

Gene regulation was examined in Caco‐2 cells. RNA stability was measured after interruption of transcription. The activity of deletion mutations of the hIGFBP‐3 promoter was examined in reporter assays. Transcription factor binding to promoter DNA was analyzed.

Results:

Butyrate did not increase the transcription of a repressor because it inhibited hIGFBP‐3 mRNA in the absence of protein synthesis. Nor did butyrate decrease the stability of hIGFBP‐3 mRNA. Analysis of the hIGFBP‐3 promoter demonstrated a butyrate‐response element that included the binding sites for p300 and Sp1/Sp3. Transfection of Caco‐2 cells with E1A, an inhibitor of p300 acetyltransferase activity, reversed the butyrate‐induced repression of hIGFBP‐3. Because Sp3 represses the initiation of transcription, we studied whether butyrate induced Sp3 acetylation. Electrophoretic mobility shift assays of nuclei extracted from Caco‐2 cells treated with 5 mmol/L butyrate demonstrated an extra, heavier band in addition to the Sp3‐DNA binding in untreated cells. This corresponded to a protein, detected only in butyrate treated cells, that was identified both by an anti‐Sp3 antibody and by an anti‐acetyl lysine antibody.

Conclusions:

This study demonstrates that butyrate increases the acetylation of a nonhistone protein, Sp3, catalyzed by p300 acetyltransferase activity.
Long Term Nutritional Rehabilitation by Gastrostomy in Israeli Patients with Cystic Fibrosis: Clinical Outcome in Advanced Pulmonary Disease
Authors: Efrati Ori , Mei‐Zahav Meir , Rivlin Joseph , Kerem Eitan , Blau Hannah , Barak Asher , Bujanover Yoram , Augarten Arie , Cochavi Brijit , Yahav Yaacov , Modan‐Moses Dalit
Published: February 2006

Objectives:

Several studies have shown a linear correlation between nutritional status and pulmonary function in patients with cystic fibrosis. Our study aims were: 1) To evaluate the effect of nutritional supplementation via gastrostomy on nutritional, clinical, and pulmonary parameters, and 2) To identify predicting factors for success of long‐term nutritional rehabilitation.

Methods:

Twenty‐one Israeli patients, aged 8 months to 20 years, underwent gastrostomy insertion from 1992 to 2001. All patients were pancreatic insufficient, and all carried severe mutations (W1282X in 62% of the patients). Anthropometric and clinical data were obtained for each patient: 0−12 months before and 6−12 months and 18−24 months after gastrostomy placement. Standard deviation scores (SDS) for height, weight, and body mass index as well as percent of height‐appropriate body weight were calculated.

Results:

The mean percent‐of‐predicted forced expiratory volume in 1 second (FEV1) decreased significantly during the first year of gastrostomy feeding (n = 16), from 44.2% ± 13.9 to 41% ± 13.3 (P = 0.05). However, during the second year of therapy (n = 10), a trend toward improvement was observed (from 39.4 ± 12.1 to 41.4 ± 16.1). Weight, and BMI z‐scores as well as weight percent‐of ideal body weight increased significantly. Height z‐score for age decreased during the first year (from −1.9 ± 1.3 to −2.1 ± 1.4), However, a trend toward improvement was observed during the second year. A significant correlation was found between the change in weight z‐score and height z‐score during the first (r = 0.488, P = 0.016) and the second (r = 0.825, P < 0.001) years. There was no difference between compliers and noncompliers regarding height, weight, and BMI either before or after gastrostomy placement. A significant correlation between age at insertion of gastrostomy and improvement in height z‐score (r = 0.52, P = 0.016) was observed. Cystic fibrosis related diabetes (n = 8) did not affect the response to supplemental feeding.

Conclusions:

We observed a trend toward improvement of pulmonary disease during the second year, and a significant improvement in weight, height, and BMI z‐scores. Compliance, diabetes, and young age prior to tube insertion did not predict success of nutritional rehabilitation.
Endoscopic Outcome Beyond Esophageal Variceal Eradication in Children with Extrahepatic Portal Venous Obstruction
Authors: Itha Srivenu , Yachha Surender Kumar
Published: February 2006

Objectives:

To find out the recurrence of esophageal varices, evolution of gastric varices, portal hypertensive gastropathy (PHG) and risk of rebleeding following esophageal variceal eradication.

Methods:

Between 1992 and 2002, children with extrahepatic portal venous obstruction (EHPVO) and bleeding from esophageal varices received endoscopic injection sclerotherapy until eradication. Surveillance endoscopy was performed initially at 3 months and subsequently at intervals of 6 months to one year to detect esophageal and gastric varices, and PHG. Gastric varices were classified as gastroesophageal (GOV) or isolated gastric varices (IGV). Gastroesophageal varices included types GOV1 and GOV2 that extend along lesser and greater curvatures respectively. Patients who had recurrence of bleeding were evaluated by emergency upper gastrointestinal endoscopy.

Results:

163 of 183 children who achieved esophageal variceal eradication were evaluated. Esophageal varices recurred in 40% cases. Primary gastric varices (before sclerotherapy) were seen in 61% cases [GOV 98% (83% GOV1, 15% GOV2) and IGV 2%] and secondary (after sclerotherapy) in 28% [GOV 71% (47% GOV1, 24% GOV2) and IGV 29%]. Secondary gastric varices were distributed as 20% GOV1, 42% GOV2 and 87% IGV. Frequency of gastric varices before sclerotherapy and at the last follow up showed decrease in GOV1 from 82 to 56 (P = 0.02), increase in GOV2 from 15 to 23 and increase in IGV from 2 to 15 (P < 0.001). PHG increased in frequency from 12% to 41% (P < 0.001) and severity from one patient to 12 (P < 0.001). Eleven cases had rebleeding from gastric varices (5 GOV1, 4 GOV2 and 2 IGV).

Conclusions:

Following esophageal variceal eradication in children with EHPVO a significant decrease in GOV1, increase in IGV and increased frequency and severity of PHG takes place. Small rebleeding risk persists from gastric varices irrespective of the type.
Nocturnal Acid Breakthrough in Children with Reflux Esophagitis Taking Proton Pump Inhibitors
Authors: Pfefferkorn Marian D , Croffie Joseph M , Gupta Sandeep K , Molleston Jean P , Eckert George J , Corkins Mark R , Fitzgerald Joseph F
Published: February 2006

Objectives:

We aimed to determine if nocturnal acid breakthrough occurs in children receiving proton pump inhibitors for reflux esophagitis, and to compare the healing of esophagitis in children with nocturnal acid breakthrough receiving proton pump inhibitors ± ranitidine.

Methods:

This is a prospective, double‐blind study. Endoscopic and histologic esophagitis were scored 0‐4 and 0‐3, respectively. Patients were treated with a proton pump inhibitor twice daily and esophagogastric pH monitoring was performed at week 3. Patients with nocturnal acid breakthrough were randomized. One group received ranitidine and the other received placebo at bedtime in addition to proton pump inhibitor therapy. Endoscopy was performed on all patients (with pH monitoring on patients with nocturnal acid breakthrough) during the 17th week of therapy.

Results:

We enrolled 18 patients, ages 1 to 13 years (mean = 10.3 years). Mean baseline endoscopic and histologic scores were 3.1 +/− 1.4 and 1.8 +/− 0.7, respectively. Mean dose of proton pump inhibitor was 1.3 mg/kg +/− 0.6. Nocturnal acid breakthrough was documented in 16/18 (89%) patients. Seven patients received ranitidine and 9 received placebo. The reflux index improved: mean of 14.3 at baseline, 2.0 at week 3 (P= 0.0001), and 5.1 at week 17 (P= 0.09). Nocturnal acid breakthrough persisted in 9/12 (75%) patients, 3 of whom received ranitidine at bedtime. Esophagitis improved inallpatients following therapy: mean endoscopy and histology scores were 1.6 +/− 1.8 (P= 0.0020) and 0.8 +/− 0.9 (P= 0.0013), respectively. Symptoms significantly improved from a mean score of 2.0 at baseline to 0.4 at week 17 (P= 0.0001).

Conclusions:

Nocturnal acid breakthrough is common in pediatric patients treated with proton pump inhibitors. Reflux index remains normal in spite of nocturnal acid breakthrough. Symptoms and esophagitis continued to improve during therapy in spite of nocturnal acid breakthrough. There appears to be no additional benefit to supplementation with ranitidine at bedtime.
Mimicking the action of GroEL in molecular dynamics simulations: Application to the refinement of protein structures
Authors: Fan Hao , Mark Alan E.
Published: February 2006

Bacterial chaperonin, GroEL, together with its co‐chaperonin, GroES, facilitates the folding of a variety of polypeptides. Experiments suggest that GroEL stimulates protein folding by multiple cycles of binding and release. Misfolded proteins first bind to an exposed hydrophobic surface on GroEL. GroES then encapsulates the substrate and triggers its release into the central cavity of the GroEL/ES complex for folding. In this work, we investigate the possibility to facilitate protein folding in molecular dynamics simulations by mimicking the effects of GroEL/ES namely, repeated binding and release, together with spatial confinement. During the binding stage, the (metastable) partially folded proteins are allowed to attach spontaneously to a hydrophobic surface within the simulation box. This destabilizes the structures, which are then transferred into a spatially confined cavity for folding. The approach has been tested by attempting to refine protein structural models generated using the ROSETTA procedure for ab initio structure prediction. Dramatic improvements in regard to the deviation of protein models from the corresponding experimental structures were observed. The results suggest that the primary effects of the GroEL/ES system can be mimicked in a simple coarse‐grained manner and be used to facilitate protein folding in molecular dynamics simulations. Furthermore, the results support the assumption that the spatial confinement in GroEL/ES assists the folding of encapsulated proteins.
A general model of G protein‐coupled receptor sequences and its application to detect remote homologs
Authors: Wistrand Markus , Käll Lukas , Sonnhammer Erik L.L.
Published: February 2006

G protein‐coupled receptors (GPCRs) constitute a large superfamily involved in various types of signal transduction pathways triggered by hormones, odorants, peptides, proteins, and other types of ligands. The superfamily is so diverse that many members lack sequence similarity, although they all span the cell membrane seven times with an extracellular N and a cytosolic C terminus. We analyzed a divergent set of GPCRs and found distinct loop length patterns and differences in amino acid composition between cytosolic loops, extracellular loops, and membrane regions. We configured GPCRHMM, a hidden Markov model, to fit those features and trained it on a large dataset representing the entire superfamily. GPCRHMM was benchmarked to profile HMMs and generic transmembrane detectors on sets of known GPCRs and non‐GPCRs. In a cross‐validation procedure, profile HMMs produced an error rate nearly twice as high as GPCRHMM. In a sensitivity‐selectivity test, GPCRHMM's sensitivity was about 15% higher than that of the best transmembrane predictors, at comparable false positive rates. We used GPCRHMM to search for novel members of the GPCR superfamily in five proteomes. All in all we detected 120 sequences that lacked annotation and are potentially novel GPCRs. Out of those 102 were found in Caenorhabditis elegans, four in human, and seven in mouse. Many predictions (65) belonged to Pfam domains of unknown function. GPCRHMM strongly rejected a family of arthropod‐specific odorant receptors believed to be GPCRs. A detailed analysis showed that these sequences are indeed very different from other GPCRs. GPCRHMM is available at http://gpcrhmm.cgb.ki.se.
The solution structure of the methylated form of the N‐terminal 16‐kDa domain of Escherichia coli Ada protein
Authors: Takinowaki Hiroto , Matsuda Yasuhiro , Yoshida Takuya , Kobayashi Yuji , Ohkubo Tadayasu
Published: February 2006

The N‐terminal 16‐kDa domain of Escherichia coli Ada protein (N‐Ada16k) repairs DNA methyl phosphotriester lesions by an irreversible methyl transfer to its cysteine residue. Upon the methylation, the sequence‐specific DNA binding affinity for the promoter region of the alkylation resistance genes is enhanced by 103‐fold. Then, it acts as a transcriptional regulator for the methylation damage. In this paper, we identified the methyl acceptor residue of N‐Ada16k and determined the solution structure of the methylated form of N‐Ada16k by using NMR and mass spectrometry. The results of a 13C‐filtered 1H‐13C HMBC experiment and MALDI‐TOF MS and MS/MS experiments clearly showed that the methyl acceptor residue is Cys38. The solution structure revealed that it has two distinct subdomains connected by a flexible linker loop: the methyltransferase (MTase) subdomain with the zinc–thiolate center, and the helical subdomain with a helix‐turn‐helix motif. Interestingly, there is no potential hydrogen bond donor around Cys38, whereas the other three cysteine residues coordinated to a zinc ion have potential donors. Hence, Cys38 could retain its inherent nucleophilicity and react with a methyl phosphotriester. Furthermore, the structure comparison shows that there is no indication of a remarkable conformational change occurring upon the methylation. This implies that the electrostatic repulsion between the negatively charged DNA and the zinc–thiolate center may avoid the contact between the MTase subdomain and the DNA in the nonmethylated form. Thus, after the Cys38 methylation, the MTase subdomain can bind the cognate DNA because the negative charge of the zinc–thiolate center is reduced.
Domain:domain interactions within Hop, the Hsp70/Hsp90 organizing protein, are required for protein stability and structure
Authors: Carrigan Patricia E. , Sikkink Laura A. , Smith David F. , Ramirez‐Alvarado Marina
Published: February 2006

The major heat shock protein (Hsp) chaperones Hsp70 and Hsp90 both bind the co‐chaperone Hop (Hsp70/Hsp90 organizing protein), which coordinates Hsp actions in folding protein substrates. Hop contains three tetratricopeptide repeat (TPR) domains that have binding sites for the conserved EEVD C termini of Hsp70 and Hsp90. Crystallographic studies have shown that EEVD interacts with positively charged amino acids in Hop TPR‐binding pockets (called carboxylate clamps), and point mutations of these carboxylate clamp positions can disrupt Hsp binding. In this report, we use circular dichroism to assess the effects of point mutations and Hsp70/Hsp90 peptide binding on Hop conformation. Our results show that Hop global conformation is destabilized by single point mutations in carboxylate clamp positions at pH 5, while the structure of individual TPR domains is unaffected. Binding of peptides corresponding to the C termini of Hsp70 and Hsp90 alters the global conformation of wild‐type Hop, whereas peptide binding does not alter conformation of individual TPR domains. These results provide biophysical evidence that Hop‐binding pockets are directly involved with domain:domain interactions, both influencing Hop global conformation and Hsp binding, and contributing to proper coordination of Hsp70 and Hsp90 interactions with protein substrates.
Octapeptide repeat insertions increase the rate of protease‐resistant prion protein formation
Authors: Moore Roger A. , Herzog Christian , Errett John , Kocisko David A. , Arnold Kevin M. , Hayes Stanley F. , Priola Suzette A.
Published: February 2006

A central feature of transmissible spongiform encephalopathies (TSE or prion diseases) involves the conversion of a normal, protease‐sensitive glycoprotein termed prion protein (PrP‐sen) into a pro‐tease‐resistant form, termed PrP‐res. The N terminus of PrP‐sen has five copies of a repeating eight amino acid sequence (octapeptide repeat). The presence of one to nine extra copies of this motif is associated with a heritable form of Creutzfeld‐Jakob disease (CJD) in humans. An increasing number of octapeptide repeats correlates with earlier CJD onset, suggesting that the rate at which PrP‐sen misfolds into PrP‐res may be influenced by these mutations. In order to determine if octapeptide repeat insertions influence the rate at which PrP‐res is formed, we used a hamster PrP amyloid‐forming peptide (residues 23–144) into which two to 10 extra octapeptide repeats were inserted. The spontaneous formation of protease‐resistant PrP amyloid from these peptides was more rapid in response to an increased number of octapeptide repeats. Furthermore, experiments using full‐length glycosylated hamster PrP‐sen demonstrated that PrP‐res formation also occurred more rapidly from PrP‐sen molecules expressing 10 extra copies of the octapeptide repeat. The rate increase for PrP‐res formation did not appear to be due to any influence of the octapeptide repeat region on PrP structure, but rather to more rapid binding between PrP molecules. Our data from both models support the hypothesis that extra octapeptide repeats in PrP increase the rate at which protease resistant PrP is formed which in turn may affect the rate of disease onset in familial forms of CJD.
Site‐specific labeling of proteins for single‐molecule FRET by combining chemical and enzymatic modification
Authors: Jäger Marcus , Nir Eyal , Weiss Shimon
Published: February 2006

An often limiting factor for studying protein folding by single‐molecule fluorescence resonance energy transfer (FRET) is the ability to site‐specifically introduce a photostable organic FRET donor (D) and a complementary acceptor (A) into a polypeptide chain. Using alternating‐laser excitation and chymotrypsin inhibitor 2 as a model, we show that chemical labeling of a unique cysteine, followed by enzymatic modification of a reactive glutamine in an N‐terminally appended substrate sequence recognition tag for transglutaminase (TGase) affords stoichiometrically D‐/A‐labeled protein suitable for single‐molecule FRET experiments. Thermodynamic data indicate that neither the presence of the TGase tag nor D/A labeling perturbs protein stability. As the N terminus in proteins is typically solvent accessible, a TGase tag can (in principle) be appended to any protein of interest by genetic engineering. Two‐step chemical/enzymatic labeling may thus represent a simple, low‐cost, and widely available strategy for D/A labeling of proteins for FRET‐based single‐molecule protein folding studies, even for non‐protein‐experts laboratories.
Methionine oxidation of monomeric λ repressor: The denatured state ensemble under nondenaturing conditions
Authors: Chugha Preeti , Sage Harvey J. , Oas Terrence G.
Published: February 2006

Although poorly understood, the properties of the denatured state ensemble are critical to the thermodynamics and the kinetics of protein folding. The most relevant conformations to cellular protein folding are the ones populated under physiological conditions. To avoid the problem of low expression that is seen with unstable variants, we used methionine oxidation to destabilize monomeric λ repressor and predominantly populate the denatured state under nondenaturing buffer conditions. The denatured ensemble populated under these conditions comprises conformations that are compact. Analytical ultracentrifugation sedimentation velocity experiments indicate a small increase in Stokes radius over that of the native state. A significant degree of α‐helical structure in these conformations is detected by far‐UV circular dichroism, and some tertiary interactions are suggested by near‐UV circular dichroism. The characteristics of the denatured state populated by methionine oxidation in nondenaturing buffer are very different from those found in chemical denaturant.
Solution structure of PcFK1, a spider peptide active against Plasmodium falciparum
Authors: Pimentel Cyril , Choi Soo‐Jin , Chagot Benjamin , Guette Catherine , Camadro Jean‐Michel , Darbon Hervé
Published: February 2006

Psalmopeotoxin I (PcFK1) is a 33‐amino‐acid residue peptide isolated from the venom of the tarantula Psalmopoeus cambridgei. It has been recently shown to possess strong antiplasmodial activity against the intra‐erythrocyte stage of Plasmodium falciparum in vitro. Although the molecular target for PcFK1 is not yet determined, this peptide does not lyse erythrocytes, is not cytotoxic to nucleated mammalian cells, and does not inhibit neuromuscular function. We investigated the structural properties of PcFK1 to help understand the unique mechanism of action of this peptide and to enhance its utility as a lead compound for rational development of new antimalarial drugs. In this paper, we have determined the three‐dimensional solution structure by 1H two‐dimensional NMR means of recombinant PcFK1, which is shown to belong to the ICK structural superfamily with structural determinants common to several neurotoxins acting as ion channels effectors.
Fluorescence resonance energy transfer and molecular modeling studies on 4′,6‐diamidino‐2‐phenylindole (DAPI) complexes with tubulin
Authors: Arbildua José J. , Brunet Juan E. , Jameson David M. , López Maribel , Nova Esteban , Lagos Rosalba , Monasterio Octavio
Published: February 2006

The goal of this work was to determine the binding properties and location of 4′,6‐diamidino‐2‐phenylindole (DAPI) complexed with tubulin. Using fluorescence anisotropy, a dissociation constant of 5.2 ± 0.4 μM for the DAPI–tubulin complex was determined, slightly lower than that for the tubulin S complex. The influence of the C‐terminal region on the binding of DAPI to tubulin was also characterized. Using FRET experiments, and assuming a κ2 value of 2/3, distances between Co2+ bound to its high‐affinity binding site and the DAPI‐binding site and 2′,3′‐O‐(trinitrophenyl)guanosine 5′‐triphosphate bound to the exchangeable nucleotide and the DAPI‐binding site were found to be 20 ± 2 Å and 43 ± 2 Å, respectively. To locate potential DAPI‐binding sites on tubulin, a molecular modeling study was carried out using the tubulin crystal structure and energy minimization calculations. The results from the FRET measurements were used to limit the possible location of DAPI in the tubulin structure. Several candidate binding sites were found and these are discussed in the context of the various properties of bound DAPI.
Temperature‐induced reversible conformational change in the first 100 residues of α‐synuclein
Authors: McNulty Brian C. , Tripathy Ashutosh , Young Gregory B. , Charlton Lisa M. , Orans Jillian , Pielak Gary J.
Published: February 2006

Natively disordered proteins are a growing class of anomalies to the structure–function paradigm. The natively disordered protein α‐synuclein is the primary component of Lewy bodies, the cellular hallmark of Parkinson's disease. We noticed a dramatic difference in dilute solution 1H‐15N Heteronuclear Single Quantum Coherence (HSQC) spectra of wild‐type α‐synuclein and two disease‐related mutants (A30P and A53T), with spectra collected at 35°C showing fewer cross‐peaks than spectra acquired at 10°C. Here, we show the change to be the result of a reversible conformational exchange linked to an increase in hydrodynamic radius and secondary structure as the temperature is raised. Combined with analytical ultracentrifugation data showing α‐synuclein to be monomeric at both temperatures, we conclude that the poor quality of the 1H‐15N HSQC spectra obtained at 35°C is due to conformational fluctuations that occur on the proton chemical shift time scale. Using a truncated variant of α‐synuclein, we show the conformational exchange occurs in the first 100 amino acids of the protein. Our data illustrate a key difference between globular and natively disordered proteins. The properties of globular proteins change little with solution conditions until they denature cooperatively, but the properties of natively disordered proteins can vary dramatically with solution conditions.
Global structural rearrangement of the cell penetrating ribonuclease colicin E3 on interaction with phospholipid membranes
Authors: Mosbahi Khédidja , Walker Daniel , James Richard , Moore Geoffrey R. , Kleanthous Colin
Published: February 2006

Nuclease type colicins and related bacteriocins possess the unprecedented ability to translocate an enzymatic polypeptide chain across the Gram‐negative cell envelope. Here we use the rRNase domain of the cytotoxic ribonuclease colicin E3 to examine the structural changes on its interaction with the membrane. Using phospholipid vesicles as model membranes we show that anionic membranes destabilize the nuclease domain of the rRNase type colicin E3. Intrinsic tryptophan fluorescence and circular dichroism show that vesicles consisting of pure DOPA act as a powerful protein denaturant toward the rRNase domain, although this interaction can be entirely prevented by the addition of salt. Binding of E3 rRNase to DOPA vesicles is an endothermic process (ΔH = 24 kcal mol−1), reflecting unfolding of the protein. Consistent with this, binding of a highly destabilized mutant of the E3 rRNase to DOPA vesicles is exothermic. With mixed vesicles containing anionic and neutral phospholipids at a ratio of 1:3, set to mimic the charge of the Escherichia coli inner membrane, destabilization of E3 rRNase is lessened, although the melting temperature of the protein at pH 7.0 is greatly reduced from 50°C to 30°C. The interaction of E3 rRNase with 1:3 DOPA:DOPC vesicles is also highly dependent on both ionic strength and temperature. We discuss these results in terms of the likely interaction of the E3 rRNase and the related E9 DNase domains with the E. coli inner membrane and their subsequent translocation to the cell cytoplasm.
Genetic selection for protein solubility enabled by the folding quality control feature of the twin‐arginine translocation pathway
Authors: Fisher Adam C. , Kim Woojin , Delisa Matthew P.
Published: February 2006

One of the most vexing problems facing structural genomics efforts and the biotechnology enterprise in general is the inability to efficiently produce functional proteins due to poor folding and insolubility. Additionally, protein misfolding and aggregation has been linked to a number of human diseases, such as Alzheimer's. Thus, a robust cellular assay that allows for direct monitoring, manipulation, and improvement of protein folding could have a profound impact. We report the development and characterization of a genetic selection for protein folding and solubility in living bacterial cells. The basis for this assay is the observation that protein transport through the bacterial twin‐arginine translocation (Tat) pathway depends on correct folding of the protein prior to transport. In this system, a test protein is expressed as a tripartite fusion between an N‐terminal Tat signal peptide and a C‐terminal TEM1 β‐lactamase reporter protein. We demonstrate that survival of Escherichia coli cells on selective medium expressing a Tat‐targeted test protein/β‐lactamase fusion correlates with the solubility of the test protein. Using this assay, we isolated solubility‐enhanced variants of the Alzheimer's Aβ42 peptide from a large combinatorial library of Aβ42 sequences, thereby confirming that our assay is a highly effective selection tool for soluble proteins. By allowing the bacterial Tat pathway to exert folding quality control on expressed target protein sequences, we have generated a powerful tool for monitoring protein folding and solubility in living cells, for molecular engineering of solubility‐enhanced proteins or for the isolation of factors and/or cellular conditions that stabilize aggregation‐prone proteins.
Exploring the interaction between the protein kinase A catalytic subunit and caveolin‐1 scaffolding domain with shotgun scanning, oligomer complementation, NMR, and docking
Authors: Levin Aron M. , Coroneus John G. , Cocco Melanie J. , Weiss Gregory A.
Published: February 2006

The techniques of phage‐displayed homolog shotgun scanning, oligomer complementation, NMR secondary structure analysis, and computational docking provide a complementary suite of tools for dissecting protein–protein interactions. Focusing these tools on the interaction between the catalytic sub‐unit of protein kinase A (PKAcat) and caveolin‐1 scaffolding domain (CSD) reveals the first structural model for the interaction. Homolog shotgun scanning varied each CSD residue as either a wild‐type or a homologous amino acid. Wild‐type to homolog ratios from 116 different homologous CSD variants identified side‐chain functional groups responsible for precise contacts with PKAcat. Structural analysis by NMR assigned an α‐helical conformation to the central residues 84– 97 of CSD. The extensive mutagenesis data and NMR secondary structure information provided constraints for developing a model for the PKAcat–CSD interaction. Addition of synthetic CSD to phage‐displayed CSD resulted in oligomer complementation, or enhanced binding to PKAcat. Together with previous experiments examining the interaction between CSD and endothelial nitric oxide synthase (eNOS), the results suggest a general oligomerization‐dependent enhancement of binding between signal transducing enzymes and caveolin‐1.
Monitoring protein aggregation during thermal unfolding in circular dichroism experiments
Authors: Benjwal Sangeeta , Verma Shikha , Röhm Klaus‐Heinrich , Gursky Olga
Published: February 2006

Thermal unfolding monitored by spectroscopy or calorimetry is widely used to determine protein stability. Equilibrium thermodynamic analysis of such unfolding is often hampered by its irreversibility, which usually results from aggregation of thermally denatured protein. In addition, heat‐induced protein misfolding and aggregation often lead to formation of amyloid‐like structures. We propose a convenient method to monitor in real time protein aggregation during thermal folding/ unfolding transition by recording turbidity or 90° light scattering data in circular dichroism (CD) spectroscopic experiments. Since the measurements of turbidity and 90° light scattering can be done simultaneously with far‐ or near‐UV CD data collection, they require no additional time or sample and can be directly correlated with the protein conformational changes monitored by CD. The results can provide useful insights into the origins of irreversible conformational changes and test the linkage between protein unfolding or misfolding and aggregation in various macromolecular systems, including globular proteins and protein–lipid complexes described in this study, as well as a wide range of amyloid‐forming proteins and peptides.
Cooperative hydrogen bond interactions in the streptavidin–biotin system
Authors: Hyre David E. , Le Trong Isolde , Merritt Ethan A. , Eccleston John F. , Green N. Michael , Stenkamp Ronald E. , Stayton Patrick S.
Published: February 2006

The thermodynamic and structural cooperativity between the Ser45– and D128–biotin hydrogen bonds was measured by calorimetric and X‐ray crystallographic studies of the S45A/D128A double mutant of streptavidin. The double mutant exhibits a binding affinity ∼2 × 107 times lower than that of wild‐type streptavidin at 25°C. The corresponding reduction in binding free energy (ΔΔG) of 10.1 kcal/mol was nearly completely due to binding enthalpy losses at this temperature. The loss of binding affinity is 11‐fold greater than that predicted by a linear combination of the single‐mutant energetic perturbations (8.7 kcal/mol), indicating that these two mutations interact cooperatively. Crystallographic characterization of the double mutant and comparison with the two single mutant structures suggest that structural rearrangements at the S45 position, when the D128 carboxylate is removed, mask the true energetic contribution of the D128–biotin interaction. Taken together, the thermodynamic and structural analyses support the conclusion that the wild‐type hydrogen bond between D128–OD and biotin–N2 is thermodynamically stronger than that between S45–OG and biotin–N1.
The Health Promotion of Aging Workers from the Perspective of Occupational Health Professionals
Authors: Naumanen Paula
Published: February 2006

ABSTRACT 
Objectives:
This study defines and describes, from the perspective of occupational health professionals, (a) the aging worker, (b) health promotion for aging workers, (3) employees' health‐promoting activities, (4) workplace health promotion factors, (5) occupational health promotion activities, (6) copartners supporting the health promotion, and (7) the impact of health promotion activities for older workers. A model is proposed, based on these issues. Design: A cross‐sectional, qualitative design was used to analyze e‐mailed essays. Content analysis of the essays yielded themes that were validated by an independent research colleague. Sample: Ten group essays from 16 occupational health professionals who had participated in the Health Promotion Project of Aging Workers in Northern Karelia, Finland were received. Results: An aging employee was defined as a mature worker. Health promotion was defined as early prevention of the influence of health hazards. Workers can improve their health through living habits, positive attitude, personal relationships, and education. Individual work arrangements, good work atmosphere, and appreciative leadership were essential factors for the health of workers. Occupational health professionals supported workers by health examinations, workplace visits, counseling, and organizing activities. Conclusions: The impacts of health promotion activities were better health, productivity, and work satisfaction.
The Science of Emotional Intelligence
Authors: Salovey Peter , Grewal Daisy
Published: February 2006

This article provides an overview of current research on emotional intelligence. Although it has been defined in many ways, we focus on the four-branch model by Mayer and Salovey (1997) , which characterizes emotional intelligence as a set of four related abilities: perceiving, using, understanding, and managing emotions. The theory provides a useful framework for studying individual differences in abilities related to processing emotional information. Despite measurement obstacles, the evidence in favor of emotional intelligence is accumulating. Emotional intelligence predicts success in important domains, among them personal and work relationships.
Prospective Memory
Authors: Einstein Gilles O. , McDaniel Mark A.
Published: February 2006

An interesting challenge for researchers who study prospective memory is to explain how people recognize environmental events as cues for actions. Whereas some theorists propose that a capacity-consuming monitoring process is the only means by which intentions can be retrieved, we argue that the cognitive system relies on multiple processes, including spontaneous processes that reflexively respond to the presence of target events. We present evidence for the existence of spontaneous retrieval processes and apply the idea of multiple processes to mixed findings on age-related decline in prospective memory.
Presurgical Language Mapping
Authors: McDermott Kathleen B. , Watson Jason M. , Ojemann Jeffrey G.
Published: February 2006

Neurosurgical procedures for tumors or intractable epilepsy are often accompanied by risk to postoperative cognitive function; surgery in the left temporal or frontal lobes, for example, can place language functions at risk. Hence, prior to tissue extraction, one common surgical goal is to attempt to identify frontal and temporal regions that should be spared in order to preserve language function. We describe how basic research on false memory for word lists has led to a novel approach for identifying such language-related regions in healthy controls. That is, rapid presentation of semantically related words (e.g., bed, rest, awake) and phonologically related words (e.g., peep, weep, heap) with instructions to attend to relations among the words elicits activity in underlying language networks. Furthermore, it is often possible to identify the neural underpinnings of these networks in an individual person in about an hour of functional magnetic resonance imaging (fMRI) scanning. We conclude by suggesting directions for future research with this lexical-processing protocol, the overarching goal being to link basic cognitive science and clinical practice.
Drug Tolerance, Drug Addiction, and Drug Anticipation
Authors: Siegel Shepard
Published: February 2006

Environmental cues associated with drugs often elicit withdrawal symptoms and relapse to drug use. Such cues also modulate drug tolerance. The contribution of drug-associated stimuli to withdrawal and tolerance is emphasized in a Pavlovian-conditioning analysis of drug administration. Conditional responses occur in the presence of cues that have been associated with the drug in the past, such as the setting in which the drug was taken. These conditional responses mediate the expression of tolerance and withdrawal symptoms. Recently, it has become apparent that internal predrug cues, as well as environmental cues, elicit pharmacological conditional responses that contribute to tolerance and withdrawal. Such internal cues include cognitive or proprioceptive cues incidental to self-administration, drug-onset cues that are experienced shortly after administration, and emotional cues. According to the conditioning analysis, addiction treatment should incorporate learning principles to extinguish the association between stimuli (environmental and internal) present at the time of drug administration and the effects of the addictive drug.
Cigarette Smoking and Panic Psychopathology
Authors: Zvolensky Michael J. , Bernstein Amit
Published: February 2006

The present paper summarizes empirical evidence suggesting that smoking and panic problems often co-occur; that smoking is a risk factor for, and may serve to maintain, panic attacks and panic disorder; and that premorbid panic-specific vulnerability variables and full-blown panic problems are related to coping-oriented smoking motives and perhaps to the maintenance of smoking behavior. An integrative model is offered to stimulate further work on this topic, followed by future directions for research.
Adaptations to Ovulation
Authors: Gangestad Steven W. , Thornhill Randy , Garver-Apgar Christine E.
Published: February 2006

In socially monogamous species in which males heavily invest in offspring, there arises an inevitable genetic conflict between partners over whether investing males become biological fathers of their partners' offspring. Humans are such a species. The ovulatory-shift hypothesis proposes that changes in women's mate preferences and sexual interests across the cycle are footprints of this conflict. When fertile (mid-cycle), women find masculine bodily and behavioral features particularly sexy and report increased attraction to men other than current partners. Men are more vigilant of partners when the latter are fertile, which may reflect evolved counteradaptations. This adaptationist hypothesis has already generated several fruitful research programs, but many questions remain.
When Photographs Create False Memories
Authors: Garry Maryanne , Gerrie Matthew P.
Published: February 2006

Photographs help people illustrate the stories of their lives and the significant stories of their society. However, photographs can do more than illustrate events; in this article, we show that photographs can distort memory for them. We describe the course of our “false-memory implantation” research, and review recent work showing that photographs can sometimes increase—while other times decrease—false memories. First, we discuss research showing that a doctored photo, showing subjects taking a completely fictitious hot-air-balloon ride, can cultivate false memories for that experience. We hypothesize that the photograph helps subjects to imagine details about the event that they later confuse with reality. Second, we show that although photographs are indeed powerful sources of influence on memory, they are not necessarily as powerful as narrative. In fact, in certain circumstances, photographs might constrain imagination. Third, we discuss research showing that true photographs can also cultivate false memories. Finally, we present recent work showing that photographs can create false memories for current events.