bluuebunny/crossref_metadata_2025_split huggingface dataset:
Identifying putative Mycobacterium tuberculosis Rv2004c protein sequences that bind specifically to U937 macrophages and A549 epithelial cells
Authors: Forero Martha , Puentes Álvaro , Cortés Jimena , Castillo Fabio , Vera Ricardo , Rodríguez Luis E. , Valbuena John , Ocampo Marisol , Curtidor Hernando , Rosas Jaiver , García Javier , Barrera Gloria , Alfonso Rosalba , Patarroyo Manuel A. , Patarroyo Manuel E. 
Published: October 2005

AbstractVirulence and immunity are still poorly understood in Mycobacterium tuberculosis. The H37Rv M. tuberculosis laboratory strain genome has been completely sequenced, and this along with proteomic technology represent powerful tools contributing toward studying the biology of target cell interaction with a facultative bacillus and designing new strategies for controlling tuberculosis. Rv2004c is a putative M. tuberculosis protein that could have specific mycobacterial functions. This study has revealed that the encoding gene is present in all mycobacterium species belonging to the M. tuberculosis complex. Rv2004c gene transcription was observed in all of this complex's strains except Mycobacterium bovis and Mycobacterium microti. Rv2004c protein expression was confirmed by using antibodies able to recognize a 54‐kDa molecule by immunoblotting, and its location was detected on the M. tuberculosis surface by transmission electron microscopy, suggesting that it is a mycobacterial surface protein. Binding assays led to recognizing high activity binding peptides (HABP); five HABPs specifically bound to U937 cells, and six specifically bound to A549 cells. HABP circular dichroism suggested that they had an α‐helical structure. HABP–target cell interaction was determined to be specific and saturable; some of them also displayed greater affinity for A549 cells than U937 cells. The critical amino acids directly involved in their interaction with U937 cells were also determined. Two probable receptor molecules were found on U937 cells and five on A549 for the two HABPs analyzed. These observations have important biological significance for studying bacillus–target cell interactions and implications for developing strategies for controlling this disease.
High‐affinity fragment complementation of a fibronectin type III domain and its application to stability enhancement
Authors: Dutta Sanjib , Batori Vincent , Koide Akiko , Koide Shohei 
Published: October 2005

AbstractThe tenth fibronectin type III (FN3) domain of human fibronectin (FNfn10), a prototype of the ubiquitous FN3 domain, is a small, monomeric β‐sandwich protein. In this study, we have bisected FNfn10 in each loop to generate a total of six fragment pairs. We found that fragment pairs bisected at multiple loops of FNfn10 show complementation in vivo as tested with a yeast two‐hybrid system. The dissociation constant of these fragment pairs determined in vitro were as low as 3 nM, resulting in one of the tightest fragment complementation systems reported so far. Furthermore, we show that the affinity of fragment complementation is correlated with the stability of the uncut parent protein. Exploring this correlation, we screened a yeast two‐hybrid library of one fragment and identified mutations that suppress the effect of a destabilizing mutation in the other fragment. One of the identified mutations significantly increased the stability of the uncut wild‐type protein, proving that fragment complementation can be used as a novel strategy for the selection of proteins with enhanced stability.
The crystal structure of the cis‐proline to glycine variant (P114G) of ribonuclease A
Authors: Schultz David A. , Friedman Alan M. , White Mark A. , Fox Robert O. 
Published: October 2005

AbstractReplacement of a cis‐proline by glycine at position 114 in ribonuclease A leads to a large decrease in thermal stability and simplifies the refolding kinetics. A crystallographic approach was used to determine whether the decrease in thermal stability results from the presence of a cis glycine peptide bond, or from a localized structural rearrangement caused by the isomerization of the mutated cis 114 peptide bond. The structure was solved at 2.0 Å resolution and refined to an R‐factor of 19.5% and an Rfree of 21.9%. The overall conformation of the protein was similar to that of wild‐type ribonuclease A; however, there was a large localized rearrangement of the mutated loop (residues 110–117—a 9.3 Å shift of the Cα atom of residue 114). The peptide bond before Gly114 is in the trans configuration. Interestingly, a large anomalous difference density was found near residue 114, and was attributed to a bound cesium ion present in the crystallization experiment. The trans isomeric configuration of the peptide bond in the folded state of this mutant is consistent with the refolding kinetics previously reported, and the associated protein conformational change provides an explanation for the decreased thermal stability.
Misfolding of a bacterial autotransporter
Authors: Mogensen Jesper E. , Kleinschmidt Jörg H. , Schmidt M. Alexander , Otzen Daniel E. 
Published: October 2005

AbstractThe adhesin involved in diffuse adherence (AIDA) is an autotransporter protein that confers the diffuse adherence phenotype to certain diarrheagenic Escherichia coli strains. It consists of a 49 amino acid signal peptide, a 797 amino acid passenger domain, and a 440 amino acid β‐domain integrated into the outer membrane. The β‐domain consists of two parts: the β1‐domain, which is predicted to form two β‐strands on the bacterial cell surface, and the β2‐domain, which constitutes the transmembrane domain. We have previously shown that the β‐domain can be folded from the urea‐denatured state when bound to a nickel column during purification. It has not been possible to achieve proper refolding of the β‐domain in solution; instead, a misfolded state C is formed. Here, we characterize this misfolded state in greater detail, showing that despite being misfolded, C can be analyzed as a conventional conformational state, with cooperative unfolding in urea and SDS as well as showing simple exponential kinetics during its formation in the presence of lipid vesicles and detergent micelles. The kinetics of formation of C is sensitive to the lipid composition in vesicles. We have also attempted to identify biological factors that might aid folding of the β‐domain to the properly folded state. However, no purified periplasmic or cytosolic chaperone was found to increase folding yields, and no factor in a periplasmic extract was identified that could bind to C. We conclude that it is the exposure to the unique spatial arrangement of the bacterial cell that leads to proper refolding of the β‐domain.
Conformational changes accompany phosphorylation of the epidermal growth factor receptor C‐terminal domain
Authors: Lee Nam Y. , Koland John G. 
Published: October 2005

AbstractThe precise regulation of epidermal growth factor receptor (EGFR) signaling is crucial to its function in cellular growth control. Various studies have suggested that the C‐terminal phosphorylation domain, itself a substrate for the EGFR kinase activity, exerts a regulatory influence upon it, although the molecular mechanism for this regulation is unknown. The fluorescence resonance energy transfer (FRET) technique was employed to examine how C‐terminal domain conformational changes in the context of receptor activation and autophosphorylation might regulate EGFR enzymatic activity. A novel FRET reporter system was devised in which recombinant purified EGFR intracellular domain (ICD) proteins of varying C‐terminal lengths were site‐specifically labeled at their extreme C termini with blue fluorescent protein (BFP) and a fluorescent nucleotide analog, 2′(3′)‐O‐(2,4,6‐trinitrophenyl)‐adenosine 5′‐triphosphate (TNP‐ATP), binding at their active sites. This novel BFP/TNP‐ATP FRET pair demonstrated efficient energy transfer as evidenced by appreciable BFP‐donor quenching by bound TNP‐ATP. In particular, a marked reduction in energy transfer was observed for the full‐length BFP‐labeled EGFR‐ICD protein upon phosphorylation, likely reflecting its movement away from the active site. The estimated distances from the BFP module to the TNP‐ATP‐occupied active site for the full‐length and C‐terminally truncated proteins also reveal the possible folding geometry of this domain with respect to the kinase core. The present studies demonstrate the first use of BFP/TNP‐ATP as a FRET reporter system. Furthermore, the results described here provide biophysical evidence for phosphorylation‐dependent conformational changes in the C‐terminal phosphorylation domain and its likely interaction with the kinase core.
Specificity and reversibility of the transpeptidation reaction catalyzed by the Streptomyces R61 D‐Ala‐D‐Ala peptidase
Authors: Rhazi Noureddine , Delmarcelle Michael , Sauvage Eric , Jacquemotte Françoise , Devriendt Kris , Tallon Valérie , Ghosez Léon , Frère Jean‐Marie 
Published: October 2005

AbstractThe specificity of the Streptomyces R61 penicillin‐sensitive D‐Ala‐D‐Ala peptidase has been re‐examined with the help of synthetic substrates. The products of the transpeptidation reactions obtained with Gly‐L‐Xaa dipeptides as acceptor substrates are themselves poor substrates of the enzyme. This is in apparent contradiction with the classically accepted specificity rules for D‐Ala‐D‐Ala peptidases. The Gly‐L‐Xaa dipeptide is regenerated by both the hydrolysis and transpeptidation reactions. The latter reaction is observed when another Gly‐L‐Xaa peptide or D‐Alanine are supplied as acceptors. Utilization of substrates in which the terminal ‐COO− group has been esterified or amidated shows that a free carboxylate is not an absolute prerequisite for activity. The results are discussed in the context of the expected reversibilty of the transpeptidation reaction.
Structure of a shark IgNAR antibody variable domain and modeling of an early‐developmental isotype
Authors: Streltsov Victor A. , Carmichael Jennifer A. , Nuttall Stewart D. 
Published: October 2005

AbstractThe new antigen receptor (IgNAR) antibodies from sharks are disulphide bonded dimers of two protein chains, each containing one variable and five constant domains. Three types of IgNAR variable domains have been discovered, with Type 3 appearing early in shark development and being overtaken by the antigen‐driven affinity‐matured Type 1 and 2 response. Here, we have determined the first structure of a naturally occurring Type 2 IgNAR variable domain, and identified the disulphide bond that links and stabilizes the CDR1 and CDR3 loops. This disulphide bridge locks the CDR3 loop in an “upright” conformation in contrast to other shark antibody structures, where a more lateral configuration is observed. Further, we sought to model the Type 3 isotype based on the crystallographic structure reported here. This modeling indicates (1) that internal Type 3‐specific residues combine to pack into a compact immunoglobulin core that supports the CDR loop regions, and (2) that despite apparent low‐sequence variability, there is sufficient plasticity in the CDR3 loop to form a conformationally diverse antigen‐binding surface.
Crystal structure of the N‐terminal domain of E. coli Lon protease
Authors: Li Mi , Rasulova Fatima , Melnikov Edward E. , Rotanova Tatyana V. , Gustchina Alla , Maurizi Michael R. , Wlodawer Alexander 
Published: October 2005

AbstractWe report here the first crystal structure of the N‐terminal domain of an A‐type Lon protease. Lon proteases are ubiquitous, multidomain, ATP‐dependent enzymes with both highly specific and non‐specific protein binding, unfolding, and degrading activities. We expressed and purified a stable, monomeric 119‐amino acid N‐terminal subdomain of the Escherichia coli A‐type Lon protease and determined its crystal structure at 2.03 Å (Protein Data Bank [PDB] code 2ANE). The structure was solved in two crystal forms, yielding 14 independent views. The domain exhibits a unique fold consisting primarily of three twisted β‐sheets and a single long α‐helix. Analysis of recent PDB depositions identified a similar fold in BPP1347 (PDB code 1ZBO), a 203‐amino acid protein of unknown function from Bordetella parapertussis, crystallized as part of a structural genomics effort. BPP1347 shares sequence homology with Lon N‐domains and with a family of other independently expressed proteins of unknown functions. We postulate that, as is the case in Lon proteases, this structural domain represents a general protein and polypeptide interaction domain.
NMR structure of the conserved hypothetical protein TM0487 from Thermotoga maritima: Implications for 216 homologous DUF59 proteins
Authors: Almeida Marcius S. , Herrmann Torsten , Peti Wolfgang , Wilson Ian A. , Wüthrich Kurt 
Published: October 2005

AbstractThe NMR structure of the conserved hypothetical protein TM0487 from Thermotoga maritima represents an α/β‐topology formed by the regular secondary structures α1–β1–β2–α2–β3–β4–α3– β5–310–α4, with a small anti‐parallel β‐sheet of β‐strands 1 and 2, and a mixed parallel/anti‐parallel β‐sheet of β‐strands 3–5. Similar folds have previously been observed in other proteins, with amino acid sequence identity as low as 3% and a variety of different functions. There are also 216 sequence homologs of TM0487, which all have the signature sequence of domains of unknown function 59 (DUF59), for which no three‐dimensional structures have as yet been reported. The TM0487 structure thus presents a platform for homology modeling of this large group of DUF59 proteins. Conserved among most of the DUF59s are 13 hydrophobic residues, which are clustered in the core of TM0487. A putative active site of TM0487 consisting of residues D20, E22, L23, T51, T52, and C55 is conserved in 98 of the 216 DUF59 sequences. Asp20 is buried within the proposed active site without any compensating positive charge, which suggests that its pKa value may be perturbed. Furthermore, the DUF59 family includes ORFs that are part of a conserved chromosomal group of proteins predicted to be involved in Fe–S cluster metabolism.
Crystal structures and proposed structural/functional classification of three protozoan proteins from the isochorismatase superfamily
Authors: Caruthers Jonathan , Zucker Frank , Worthey Elizabeth , Myler Peter J. , Buckner Fred , Van Voorhuis Wes , Mehlin Chris , Boni Erica , Feist Tiffany , Luft Joseph , Gulde Stacey , Lauricella Angela , Kaluzhniy Oleksandr , Anderson Lori , Le Trong Isolde , Holmes Margaret A. , Earnest Thomas , Soltis Michael , Hodgson Keith O. , Hol Wim G.J. , Merritt Ethan A. 
Published: October 2005

AbstractWe have determined the crystal structures of three homologous proteins from the pathogenic protozoans Leishmania donovani, Leishmania major, and Trypanosoma cruzi. We propose that these proteins represent a new subfamily within the isochorismatase superfamily (CDD classification cd004310). Their overall fold and key active site residues are structurally homologous both to the biochemically well‐characterized N‐carbamoylsarcosine‐amidohydrolase, a cysteine hydrolase, and to the phenazine biosynthesis protein PHZD (isochorismase), an aspartyl hydrolase. All three proteins are annotated as mitochondrial‐associated ribonuclease Mar1, based on a previous characterization of the homologous protein from L. tarentolae. This would constitute a new enzymatic activity for this structural superfamily, but this is not strongly supported by the observed structures. In these protozoan proteins, the extended active site is formed by inter‐subunit association within a tetramer, which implies a distinct evolutionary history and substrate specificity from the previously characterized members of the isochorismatase superfamily. The characterization of the active site is supported crystallographically by the presence of an unidentified ligand bound at the active site cysteine of the T. cruzi structure.
Novel sequence insertion in a Mâori patient with transfusion‐dependent β‐thalassaemia
Authors: Blacklock Hilary A. , Case Jacqui , Chan Tim , Raizis Tony , Doocey Richard , Fellowes Andrew , Royle Gordon , Jackson Sharon , Brennan Stephen , George Peter 
Published: October 2005

SummaryAlthough β‐thalassaemia is common throughout the world, it has not been previously described in Polynesia. We report a novel sequence insertion where homozygosity for the defect results in transfusion‐dependent anaemia. The repeated 45 base pair (bp) insertion causes duplication of the start codon and consequent transcription from the original initiation code would be predicted to lead to the production of an irrelevant seven‐residue peptide, while residual translation from the novel initiation site would result in diminished yields of β‐globin and consequent clinical β+‐thalassaemia.
The development of esters in filtered and unfiltered wines that have been aged in oak barrels
Authors: Moreno Nerea Jiménez , Azpilicueta Carmen Ancín 
Published: October 2005

SummaryEsters are quite important for the aroma of aged red wine as they contribute to the fruity complex nature of this wine. This research paper measured the development of esters during maturation of Merlot red wines. These wines had different amounts of turbidity. The wine was aged for 18 months either after filtration with diatomaceous earth or without filtration. Both American and French oak barrel were used for storage. The results obtained showed that the unfiltered wine had a greater concentration of esters at the end of the storage period than the filtered wine. Fatty acid and ethanol esters decreased more than isoamyl acetate, independently of filtration. Ethyl octanoate and ethyl decanoate decreased both in filtered and unfiltered wine. Isoamyl acetate was formed in unfiltered wine and ethyl butyrate and ethyl hexanoate decreased in filtered wine while they remained constant in unfiltered wine.
Hierarchical analysis of population genetic structure in the monogynous ant Cataglyphis cursor using microsatellite and mitochondrial DNA markers
Authors: CLÉMENCET J. , VIGINIER B. , DOUMS C. 
Published: October 2005

AbstractDespite having winged queens, female dispersal in the monogynous ant Cataglyphis cursor is likely to be restricted because colonies reproduce by fission. We investigated the pattern of population genetic structure of this species using eight microsatellite markers and a mitochondrial DNA (mtDNA) sequence, in order to examine the extent of female and nuclear gene flow in two types of habitat. Sampling was carried out at a large spatial scale (16 sites from 2.5 to 120 km apart) as well as at a fine spatial scale (two 4.5‐km transects, one in each habitat type). The strong spatial clustering of mtDNA observed at the fine spatial scale strongly supported a restricted effective female dispersal. In agreement, patterns of the mtDNA haplotypes observed at large and fine spatial scales suggested that new sites are colonized by nearby sites. Isolation by distance and significant nuclear genetic structure have been detected at all the spatial scales investigated. The level of local genetic differentiation for mitochondrial marker was 15 times higher than for the nuclear markers, suggesting differences in dispersal pattern between the two sexes. However, male gene flow was not sufficient to prevent significant nuclear genetic differentiation even at short distances (500 m). Isolation‐by‐distance patterns differed between the two habitat types, with a linear decrease of genetic similarities with distance observed only in the more continuous of the two habitats. Finally, despite these low dispersal capacities and the potential use of parthenogenesis to produce new queens, no signs of reduction of nuclear genetic diversity was detected in C. cursor populations.
‘In vivo’ time course of plasma myeloperoxidase levels after granulocyte colony‐stimulating factor‐induced stem cell mobilization
Authors: Morabito F. , Tomaino A. , Cristani M. , Martino M. , Minciullo P. L. , Saija A. , Gangemi S. 
Published: October 2005

summary.  Administration of granulocyte colony‐stimulating factor (G‐CSF) is widely used for harvesting an adequate number of CD34+ stem cells by leukapheresis in normal donors. G‐CSF is the most established agent for the mobilization of stem cells in current clinical practice, because it has been proven to be superior to any other agent tested to date in terms of not only mobilization capacity, but also of tolerance. However, although regulatory and accrediting agencies have provided guidelines to protect donors, the short‐ and long‐term side effects of G‐CSF need to be further studied.In this study, we evaluated the time course of plasma myeloperoxidase (MPO) levels measured in a group of donors given recombinant human G‐CSF (rHuG‐CSF) at different intervals: (i) before starting rHuG‐CSF administration, (ii) on day 5 of rHuG‐CSF administration, (iii) on the same day soon after the end of the first leukapheresis procedure and (iv) 1 week after rHuG‐CSF withdrawal.Plasma MPO levels significantly increased in the donors after 5 days of rHuG‐CSF treatment, returning to the baseline values within 7 days following rHuG‐CSF withdrawal.These findings may contribute to a better understanding of G‐CSF safety profile in stem cell donors.
hDAF porcine cardiac xenograft maintains cardiac output after orthotopic transplantation into baboon – a perioperative study
Authors: Bauer Andreas , Baschnegger Heiko , Abicht Jan M. , Brandl Ulrike , Brenner Paolo , Thein Eckart , Hammer Claus , Reichart Bruno , Peter Klaus , Schmoeckel Michael , Christ Frank 
Published: October 2005

Abstract:  Background:  Only limited data are available on the physiological functional compatibility of cardiac xenografts after orthotopic pig to baboon transplantation (oXHTx). Thus we investigated hemodynamic parameters including cardiac output (CO) before and after oXHTx.Methods:  Orthotopic xenogeneic heart transplantation from nine hDAF transgeneic piglets to baboons was performed. We used femoral arterial thermodilution for the invasive assessment of CO and stroke volume.Results:  Baseline CO of the baboons after induction of anesthesia was 1.36 (1.0–1.9) l/min. 30 to 60 min after termination of the cardiopulmonary bypass, CO of the cardiac xenograft was significantly increased to 1.72 (1.3–2.1) l/min (P < 0.01). The stroke volumes of the baboon heart before transplantation and the cardiac xenograft was comparable [14.9 (11–26) vs. 11.8 (10–23) ml]. Thus the higher CO was achieved by an increase in heart rate after oXHTx [75.0 (69–110) vs. 140.0 (77–180)/min; P < 0.01]. Despite the increased CO, oxygen delivery was reduced [256 (251–354) vs. 227 (172–477) ml/min; P < 0.01] due to the inevitable hemodilution during the cardiopulmonary bypass and the blood loss caused by the surgical procedures.Conclusion:  Our results demonstrate that in the early phase after orthotopic transplantation of hDAF pig hearts to baboons, cardiac function of the donor heart is maintained and exceeds baseline CO. However, in the early intraoperative phase this was only possible by using inotropic substances and vasopressors due to the inevitable blood loss and dilution by the priming of the bypass circuit.