Top PDF Compact conformations of human protein disulfide isomerase.

Compact conformations of human protein disulfide isomerase.

Compact conformations of human protein disulfide isomerase.

between the two active sites in domains a and a’ was determined to be 40.3 A ˚ in the oxidized state and 27.6 A˚ in the reduced state. The minimum distance between these two active sites so far reported was 16 A ˚ using crosslinking technique [27], still too far for electron transfer. In the present simulations we unexpectedly observed that this distance can decrease to as low as 5.4 A ˚ (Figure 6A and Movie S1). Thus we tried to capture the intermediates in the proposed disulfide exchange reaction between domains a and a’ by using three trapping mutants, C56A, C400A and C56A/C400A. These mutants showed the same mobility on reducing SDS-PAGE, but under non-reducing conditions a thick extra band appeared solely in C56A/C400A (Figure 6B), which was expected to be the intra-molecular Cys 53 -Cys 397 disulfide- bonded molecule. The extra band was identified by mass spectrometry following tryptic digestion. A signal with a mass (m/z) of 3562.6 (Figure 6C) was detected and perfectly matched with the disulfide-linked Tyr 43 -Lys 57 and Asn 387 -Lys 401 peptide (exactly the same mass with the sum of the two peptides minus 2 Da). Under the reducing conditions this signal disappeared but with two new peptide signals instead. The m/z signals at 1853.9 and 1824.9 were 57 Da larger than the mass of Tyr 43 -Lys 57 and Asn 387 -Lys 401 respectively, due to cysteine alkylation. The above mass spectrometry data clearly demonstrates the existence of a disulfide between Cys 53 and Cys 397 in the C56A/C400A mutant, which provides prima facie evidence that domains a and a’ in an hPDI molecule can move close enough to facilitate electron transfer.
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Substrate-induced unfolding of protein disulfide isomerase displaces the cholera toxin A1 subunit from its holotoxin.

Substrate-induced unfolding of protein disulfide isomerase displaces the cholera toxin A1 subunit from its holotoxin.

Protein disulfide isomerase (PDI) is a member of the thioredoxin superfamily with an abb’xa’c structural organization that consists of two catalytic domains (a & a9) separated by two non-catalytic domains (b & b9) and an short x linker, along with an acidic C- terminal c extension [1–3]. It is mainly located in the endoplasmic reticulum (ER) where it exhibits linked but independent oxidore- ductase and chaperone activities. These activities allow it to facilitate the proper folding of nascent secretory proteins as well as the disposal of terminally misfolded proteins through the quality control mechanism of ER-associated degradation (ERAD). The structure and function of PDI is regulated by its redox status: it is a dynamic, flexible molecule which assumes a compact conforma- tion in the reduced state and a more open conformation in the oxidized state [4–6]. PDI thus acts as a redox-dependent chaperone in its interactions with certain substrate proteins [7–10]. The chaperone function of PDI is defined by its ability to prevent the aggregation of misfolded proteins [11–14]. The importance of this activity is highlighted by the link between PDI dysfunction and neurodegeneration: a S-nitrosylated form of PDI that cannot prevent protein aggregation is found in the brains of individuals with Parkinson’s or Alzheimer’s disease [15]. PDI also prevents the aggregation of a-synuclein which occurs in
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Papel da proteína dissulfeto isomerase na sinalização redox em células endoteliais...

Papel da proteína dissulfeto isomerase na sinalização redox em células endoteliais...

Protein disulfide isomerase (PDI), an oxidoreductase of endoplasmic reticulum, has emerged as a key player in cell signaling. In vascular smooth muscle cell (VSMC) PDI regulates NADPH expression and activity. Additionally, in endothelial cells Erp46, a PDI family member, is preferentially expressed. However the role of these proteins in endothelial cell or VSMC function is still unkown. The aim of the present study was to investigate the role of PDI in redox signaling induced by TNF- α in endothelial cells and by Ang II in VSMCs. In human umbilical vein endothelial cells (HUVECs) ERp46 or PDI inhibition reduced specifically TNF- α-induced ERK1/2 phosphorylation, possibly via redox modifications in Ras GTPase. ERK activation influences gene expression by the transcriptional factor AP-1 which controls MMP-9 and cathepsin B, two proteases required for angiogenesis. ERp46 or PDI silencing decreased TNF- α induced MMP-9 and cathepsin B mRNA expression, and the formation of endothelial cell sprouts in the spheroid outgrowth assay, used as a model of angiogenesis. Our study establishes that ERp46 and PDI are required for TNF- α-induced angiogenesis. In VSMCs from resistance arteries, gain/loss of function experiments showed that Nox1 expression was regulated by PDI. In SHR cells, PDI and Nox1 expression were higher as compared to wistar cells. PDI inhibition reduced Ang II-induced ROS generation, c-Src and ERK 1/2 phosphorylation, [Ca2+]i influx and mesenteric vasoconstriction in Wistar and SHR rats. Our results suggest that PDI positively regulates Nox1 dependent signaling and expression in VSMCs from resistance arteries and be could be a new player in the oxidative stress and vascular dysfunction observed in hypertension. Altogether, the results provide evidence for a role for PDI in cardiovascular pathophysiology.
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Soluble prokaryotic overexpression and purification of bioactive human granulocyte colony-stimulating factor by maltose binding protein and protein disulfide isomerase.

Soluble prokaryotic overexpression and purification of bioactive human granulocyte colony-stimulating factor by maltose binding protein and protein disulfide isomerase.

In this study, several new methods of overexpressing soluble hGCSF in the cytoplasm of E. coli were investigated, enabling efficient production of biologically active protein. The following seven N-terminal fusion tags were used: hexahistidine (His6), thioredoxin (Trx), glutathione S-transferase (GST), MBP, N- utilization substance protein A (NusA), protein disulfide bond isomerase (PDI), and the b’a’ domain of PDI (PDIb’a’). The MBP, NusA, PDI, and PDIb’a’ tags increased the solubility of hGCSF markedly at 30 uC. Lowering the expression temperature to 18uC also increased the solubility of Trx- and GST-tagged hGCSF, whereas His6-hGCSF was insoluble at both temperatures. The expression level and the solubility of the tag-fused hGCSFs were also tested in the E. coli Origami 2(DE3) strain that have mutations in both the thioredoxin reductase (trxB) and glutathione reductase (gor) genes, which may assist the disulfide bond formation in the cytoplasm of E. coli [28–30]. Simple methods of purifying hGCSF from the PDIb’a’ or MBP tagged proteins were developed using conventional chromatographic techniques. In total, 11.3 mg of biologically active hGCSF was obtained from 500 mL of culture. Silver staining indicated that the extracted hGCSF was highly pure and the endotoxin level was very low. The activity of the purified protein was measured using a bioassay with mouse M- NFS-60 myelogenous leukemia cells.
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Mapping Soluble Guanylyl Cyclase and Protein Disulfide Isomerase Regions of Interaction.

Mapping Soluble Guanylyl Cyclase and Protein Disulfide Isomerase Regions of Interaction.

There is no full-length crystal structure or model of sGC available. In addition, the most recent PDB entry of the catalytic domain of sGC (PDB: 4NI2) [19] has both α and β subunits trun- cated at C-termini. Coincidentally, these truncated regions are where the cross-links between sGC (at α Lys672 and β Lys615) and PDI occur. Thus, we employed computational modeling to extend the available crystal structure of the sGC catalytic domain and then used it for dock- ing with PDI molecules. First, the missing C-termini amino acid residues, α G663-D690 and β T609-D619 were added to the available X-ray structure of the catalytic domain of sGC (PDB 4NI2, α 471–662, β 411–608). Because the structural organization of these flexible tails are not known, we used extended conformations of peptides and then performed molecular dynamics (MD) simulations to capture the random re-organization of the added residues. Four indepen- dent MD models were produced after 100ns simulation. The visual analysis of the models revealed that the possible closest distance between sGC residues α Lys672 and β Lys615 is 61.3Å, which is twice more than the required distance to form a contact with both Lys370 and Lys409 residues of one molecule of PDI with linkers (30Å), Fig 5A. Therefore, it is not possible
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Evaluation of Leishmania donovani protein disulfide isomerase as a potential immunogenic protein/vaccine candidate against visceral Leishmaniasis.

Evaluation of Leishmania donovani protein disulfide isomerase as a potential immunogenic protein/vaccine candidate against visceral Leishmaniasis.

PDIs are essential enzymes that catalyze thiol-disulfide inter- change, ensuring the proper folding and conformation of proteins, acting as co-receptors of cell reorganization, and preventing cell toxicity associated with ER stress and protein misfolding [31,32]. PDIs may also be involved in the host mucosal immune system, inducing secretory IgA [33,34]. Although much has been learned about PDIs in higher eukaryotes, limited information is available regarding PDIs in pathogens that are important for human infections. The protein has been reported to have role in Leishmania virulence and survival [35]. The characterization of these virulence factors obviously has important implications for the design of new drugs or vaccines against Leishmania parasites. The present study deals with the cloning, expression and purification as well as molecular characterization of LdPDI. The protein was further examined for the first time for its ability to 1) stimulate the immune responses in leishmania infected cured/endemic contact individuals’ PBMCs, 2) modulate the immune response in cured hamsters infected with L. donovani and 3) protect the hamsters against L. donovani challenge when administered as DNA vaccine. The hamster (Mesocricetus auratus) has been proven to be the most appropriate experimental model as it largely reflects the clinic- pathological features of progressive human VL, including a relentless increase in visceral parasitic burden, cachexia, hepato- splenomegaly, pancytopenia, hypergammaglobulinemia and ulti- mately death. Of late, it has also been used extensively for vaccination studies [36,37].
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On The Impact Of Pressure Drop On Human Body With Mathematical Models

On The Impact Of Pressure Drop On Human Body With Mathematical Models

Where u(r) is the velocity in the axial direction,  and  are the density and viscosity of blood,  is the couple stress parameter,  is the electrical conductivity, B 0 is the external magnetic field and r is the radial coordinate.

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Hyperthermophile protein behavior: partially-structured conformations of Pyrococcus furiosus rubredoxin monomers generated through forced cold-denaturation and refolding.

Hyperthermophile protein behavior: partially-structured conformations of Pyrococcus furiosus rubredoxin monomers generated through forced cold-denaturation and refolding.

PfuTIM has a 225 residues-long polypeptide chain, and is associated into a tetrameric quaternary structure [2]. To explore whether the unusual conformational behavior seen with PfuTIM can also be observed in a much smaller protein of hyperthermo- phile evolutionary origin, we carried out the same treatment on a very small protein, the well-known and widely studied 53 residues- long, iron-sulphur cluster-containing protein, rubredoxin (PfRd) from P. furiosus [3], [4]. PfRd is one of the most kinetically thermostable proteins known to man [5],[6], with an unfolding rate of 1.9610 26 s 21 , at pH 7.0 and 100 uC. Several excellent studies exploring the thermal stability of PfRd and, in particular, its kinetic thermal stability have been described in the literature [5],[6],[7],[8],[9]. Another thing that caused us to study this protein is the fact that it contains a disproportionately large number of charged residues. Of the 53 amino acid residues in PfRd, 18 are charged residues (34%) representing only 3 of the 20 naturally-occurring amino acids (5 lysines, 7 aspartates, and 6 glutamates) which should have normally had a representation of only 15% assuming proportional representation of each amino acid. Clearly, charged residues are playing an important role in PfRd. Since we reported that in PfuTIM the observed effects owe primarily to surface salt-bridges, it became imperative that we examine a small hyperthermophile protein like PfRd, with disproportionately high numbers of charged residues. Further- more, what made PfRd interesting was the fact that it contains a small iron-sulfur cluster and also a tight aromatic cluster in its interior which we believe (based on unpublished observations which have already been communicated for publication), to be crucially linked to iron atom binding and formation of the correct native structure.
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Human Capital and the Recent Fall of Earnings Inequality in Brazil

Human Capital and the Recent Fall of Earnings Inequality in Brazil

where is a set of linear restrictions that transforms the unrestricted model (1) on restricted model (2). 8 In our case, the restriction implies that the age, trend and (orthogonal) time dummies are sufficient to explain the behavior of each estimated statistic order across cells and over time. Imposing the restrictions means estimating weighted least squares regressions on the grouped data, for each quantile and education group separately. This procedure will give us consistent estimates of . Under the null that the restrictions are valid, the minimized value follows a chi-square distribution with degrees of freedom equal to the number of restrictions. In order to construct the test statistics, we only have to sum up the weighted squared residuals, that is, the estimated percentiles minus the predicted values minus the orthogonal time dummies.
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Computational analysis of HIV-1 resistance based on gene expression profiles and the virus-host interaction network.

Computational analysis of HIV-1 resistance based on gene expression profiles and the virus-host interaction network.

studied. CBL (murine Cas-Br-M) was first identified as part of a transforming retrovirus that induces mouse pre-B and pro-B cell lymphomas [30]. CBL positively regulates receptor protein- tyrosine kinase ubiquitination as an adaptor dependent upon its variant SH2 and RING finger domains [31]. HIV nef modifies T cell signaling by enhancing CBL phosphorylation in the absence of T cell receptor engagement and co-stimulation [32]. Nef-mediated lipid raft exclusion inhibits CBL activity, which positively regulates signaling in T cells [33]. In our study, CBL was on 19,859 of the shortest paths, which suggests that it is important for HIV infection. The second gene on the list of 29 genes, IL2, is a secreted cytokine that regulates CD4+ T cell production and survival [34]. IL2 is also known to increase CD4 cell counts in HIV-infected patients [35]. Here, IL2 was implicated in the control of 12,939 virus target communication paths, contributing to the hypothesis that IL2 is involved in HIV entry. The third gene on the list of 29 genes, ABL1 (c-abl oncogene 1, non-receptor tyrosine kinase), encodes a tyrosine kinase involved in cell differentiation, cell division, cell adhesion, and stress responses [36,37,38]. ABL1 is negatively regulated by its SH3 domain, and deletion of the SH3 domain changes ABL1 into an oncogene [37]. The DNA-binding activity of ABL1 is regulated by CDC2- mediated phosphorylation [39]. ABL1 was on 7516 virus target communication paths in this study. This is the first evidence of a link between ABL1 and HIV infection.
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Human erythrocyte acetylcholinesterase in health and disease

Human erythrocyte acetylcholinesterase in health and disease

Large amounts of AChE are present in erythrocyte membrane exovesicles [32,33]. A biochemistry laboratory practice for students, based on phthalate gradient concentrations was described using erythrocyte suspensions to obtain aggregates of exovesicles, which are characterized by AChE enzyme activity, protein, and phospholipids coloration processes [33]. This laboratory experimentation matches objectives with teaching and learning activities in biochemistry courses [33]. The same experimental protocol when used in erythrocyte suspensions under the presence of the membrane fluidity fluorescent probes (diphenylhexatriene) DPH or trimethylamino-dipheny-lhexatriene (TMA-DPH) or heptadecyl-hydroxycoumarin (C17-HC) showed the presence of exovesicles in all the supernatants, confirmed by the AChE enzyme activity that is higher in those incubated with the hydrophobic probe DPH in relation to the others amphiphilic TMA-DPH and C17-HC [34]. At variance, the intensity and fluorescence anisotropy is higher in TMA-DPH and C17-HC supernatants vesicles in relation to those obtained from the DPH [34]. Therefore, amphiphilic probes TMA-DPH and C17-HC are preferentially incorporated in the exovesicles when compared with DPH [34]. The exovesiculation process can, for example, occurs during the transformation of discocyte to stomatoscyte shape in the presence of certain types of therapeutic drugs and oxidative stress [35,36]. Mathematical modeling applied to those switch RBC forms can contribute to understanding, in patients under drug therapy, the biorheogical behavior of RBCs in tissue oxygenation [37]. RBCs’ AChE enzyme activity and osmotic fragility assessment are valuable for testing the cytotoxicity degree of lipophilic drugs that may disrupt cellular membranes beyond their final usefulness, for example, as a cell nucleus target [6].
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Expressão da proteína L1 do capsídio de HPV-16 em leveduras metilotróficas

Expressão da proteína L1 do capsídio de HPV-16 em leveduras metilotróficas

VILLA, L.L.; COSTA, R.L.; PETTA, C.A.; ANDRADE, R.P.; AULT, K.A.; GIULIANO, A.R.; WHEELER, C.M.; KOUTSKY, L.A.; MALM, C.; LEHTINEN, M.; SKJELDESTAD, F.E.; OLSSON, S.E.; STEINWALL, M.; BROWN, D.R.; KURMAN, R.J.; RONNETT, B.M.; STOLER, M.H.; FERENCZY, A.; HARPER, D.M.; TAMMS, G.M.; YU, J.; LUPINACCI, L.; RAILKAR, R.; TADDEO, F.J.; JANSEN, K.U.; ESSER, M.T.; SINGS, H.L.; SAAH, A.J.; BARR, E. Prophylactic quadrivalent human papillomavirus (types 6, 11, 16, and 18) L1 virus-like particle vaccine in young women: a randomised double-blind placebo-controlled multicentre phase II efficacy trial. The Lancet Oncology v. 6, n. 5, p. 271 - 278, 2005.
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Wiki-pi: a web-server of annotated human protein-protein interactions to aid in discovery of protein function.

Wiki-pi: a web-server of annotated human protein-protein interactions to aid in discovery of protein function.

Wiki-Pi is especially useful today, as several genome-wide association studies (GWAS) are being published. GWAS studies are unbiased by current scientific knowledge (i.e. they do not have literature-bias) and often implicate genes with currently unknown biological functions to be associated with the disease under study. The number of GWAS studies has increased rapidly in the past couple of years. So far, 1,309 publications have reported GWAS results on 674 traits or diseases (www.genome.gov/gwastudies [28], accessed 2012-July-17). Though extensive work is being carried out to identify the common genetic variants that influence various diseases or traits through GWAS, the role of these genes and the exact mechanism of their action are yet to be discovered. Very little information is available about some of the GWAS- identified genes in terms of their molecular function and biological process. Wiki-Pi enables researching each of these genes and provides novel insights that may not otherwise materialize except when a scientist knows all the multiple specialized domains involved.
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Conformational Sampling and Binding Site Assessment of Suppression of Tumorigenicity 2 Ectodomain.

Conformational Sampling and Binding Site Assessment of Suppression of Tumorigenicity 2 Ectodomain.

The first seven PCs were then used in the mean shift cluster analysis provided in the Scikit- Learn program[43]. Mean shift cluster analysis is a nonparametric clustering method[44–46]. Unlike k-mean clustering method, the mean shift clustering requires no prior knowledge of the number of clusters in the multidimensional data. The algorithm classifies data points by mov- ing the data point via the density gradient defined by a kernel density estimator function with an assigned bandwidth until it reaches the maximum of a local density distribution called mode in pattern recognition. In our application here, each mode is the center of a cluster group. To perform the mean shift cluster analysis, snapshots of ST2 ECD conformations at every 10 ps from the aMD simulations were included. The quantile value was varied from 0.01 to 0.20 in the bandwidth estimation to analyze the stability of the numbers of cluster groups. Con- formations closest to the centroid of each cluster group were selected to represent each cluster and denoted as representative conformations.
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Uma resposta parcial para a conjectura CPE, estimativas de diâmetro e variedades com energia constante

Uma resposta parcial para a conjectura CPE, estimativas de diâmetro e variedades com energia constante

In the last part, after introducing the concept of manifold with constant en- ergy, we proved that the sphere and the flat torus are the only compact surfaces with constant energy. In higher dimensions though, simple examples show that this is not true in general, but it is if we assume that the Ricci tensor is bigger than the one of the corresponding sphere. Since surface are Einstein manifolds, it seems to be natural to try to generalize Theorem 6.1 under Einstein assumption. Even in dimension four, this may be an interesting problem.

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Looking at tricellulin expression in the brain

Looking at tricellulin expression in the brain

For western blot analysis, total cell extracts were obtained in ice-cold 1x Cell Lysis Buffer plus 1nM phenylmenthylsulfonil fluoride (PMSF) and sonicated (Ultrasonic Processor UP100H, Hielscher-Ultrasound Technology, Teltow, Germany). The lysates were centrifuged at 14000g for 10 minutes, at 4 ºC, and the supernatants were collected and stored at -80 ºC. Protein concentrations were determined using microplate reader (PR 2100 Microplate Reader, Bio-Rad Laboratories). Proteins were denatured by boiling for 5 min in sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) sample buffer (0.125 M Tris- HCl, 40% glycerol, 4% SDS, 1% β-mercaptoethanol and 0.2% bromo-phenol blue). Equal amounts of protein sample were separated on a polyacrylamide gel and transferred to a nitrocellulose membrane. Following transfer and blocking with a 5% milk solution, the blots were incubated overnight at 4 ºC with the primary antibodies rabbit anti-tricellulin (1:500) or mouse anti-β-actin (1:5000) diluted in blocking solution. After three washes with Tween20- Tris buffered saline (T-TBS), the membranes were incubated with HorseRadish Peroxidase (HRP)-conjugated anti-rabbit or anti-mouse secondary antibodies for 1 hour at RT. Secondary antibodies were diluted in 5% BSA in T-TBS solution. After three washes with T- TBS, chemiluminescent detection was performed by LumiGLO® and bands were visualized by autoradiography with Hyperfilm ECL. The intensity of the bands was quantified by scanning densitometry, standardized with respect to -actin and the protein levels were expressed as fold change.
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Ablation of Prion Protein in Wild Type Human Amyloid Precursor Protein (APP) Transgenic Mice Does Not Alter The Proteolysis of APP, Levels of Amyloid-β or Pathologic Phenotype.

Ablation of Prion Protein in Wild Type Human Amyloid Precursor Protein (APP) Transgenic Mice Does Not Alter The Proteolysis of APP, Levels of Amyloid-β or Pathologic Phenotype.

Alzheimer’s disease (AD) is the most common form of dementia affecting 30 million individu- als world-wide [1,2]. Age is the greatest risk factor for AD, with the incidence doubling every 5 years after age 65. Therefore, with our ageing population, AD is placing immense financial and social pressure on society. Currently there are no treatments that either cure or halt the pro- gression of this neurodegenerative disease [3]. The majority (>95%) of AD cases have no underlying genetic mutation and are referred to as sporadic or late-onset AD [4]. In a small proportion of cases, mutations in the genes encoding the amyloid precursor protein (APP) or presenilin (PS) 1 or PS2 give rise to early onset, familial AD [4]. The disease is characterised by the deposition in the brain of extracellular plaques of amyloid-β (Aβ) which is derived from the proteolytic processing of APP [5], along with intracellular neurofibrillary tangles of
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Itm2a is a Pax3 target gene, expressed at sites of skeletal muscle formation in vivo.

Itm2a is a Pax3 target gene, expressed at sites of skeletal muscle formation in vivo.

The paired-box homeodomain transcription factor Pax3 is a key regulator of the nervous system, neural crest and skeletal muscle development. Despite the important role of this transcription factor, very few direct target genes have been characterized. We show that Itm2a, which encodes a type 2 transmembrane protein, is a direct Pax3 target in vivo, by combining genetic approaches and in vivo chromatin immunoprecipitation assays. We have generated a conditional mutant allele for Itm2a, which is an imprinted gene, by flanking exons 2–4 with loxP sites and inserting an IRESnLacZ reporter in the 39 UTR of the gene. The LacZ reporter reproduces the expression profile of Itm2a, and allowed us to further characterize its expression at sites of myogenesis, in the dermomyotome and myotome of somites, and in limb buds, in the mouse embryo. We further show that Itm2a is not only expressed in adult muscle fibres but also in the satellite cells responsible for regeneration. Itm2a mutant mice are viable and fertile with no overt phenotype during skeletal muscle formation or regeneration. Potential compensatory mechanisms are discussed.
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Improving the measurement of semantic similarity between gene ontology terms and gene products: insights from an edge- and IC-based hybrid method.

Improving the measurement of semantic similarity between gene ontology terms and gene products: insights from an edge- and IC-based hybrid method.

Two commonly used pairwise strategies, the maximum and best-match average, were chosen based on node- and edge-based semantic similarity methods to evaluate the functional similarity between two gene products. In general, the BMA approach was a better choice than MAX for estimating the functional conservation of orthologs (Table 3), and evaluating correlations with sequence, domain and ECC similarities (Figure 5, Figure S7). The MAX strategy which considers the best match among all term pairs of two gene products, could be potentially adversely affected by incorrect annotations or the noise along with the IEA annotations [33]. This may explain why HRSS (MAX) showed poor and sometimes unstable performance including IEA annotations in the evaluation based on human-mouse orthologs (Figure S4A and Table S7) and CESSM platform (Figure S6A and B in MF ontology). The MAX strategy could assess if two gene products share a similar function, but underestimates their dissimilarity. As Figure 3. Statistical significance of system for evaluating the functional similarity (BMA) of human-mouse orthologs. The evaluation system was based for the BP, CC and MF ontologies (A) including or (B) excluding IEA annotations. The histograms (measured on the left y-axis) indicate the mean and standard error of the functional similarities of observed orthologs, and the lines (measured on the right y-axis) show the Z- score values calculated from the ASVs of observed orthologous pairs and randomized pairs (see Materials and Methods). HRSS achieved the highest Z-score in all cases. Error bars indicate one standard error.
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Expression, purification and molecular analysis of the human ZNF706 protein

Expression, purification and molecular analysis of the human ZNF706 protein

The sequence alignment between HsZNF706 and the zinc-finger region of human Zinc- Fingers and Homeo- boxes 1 (ZHX1) (PDB code 2GHF) showed 34.1% iden- tity, with 41 amino acids aligned. HsZNF706 has 76 amino acid residues in the sequence, and the 2GHF template has 102 residues. Nine residues from the N- terminus of the target were removed. The structure ge- nerated by homology modeling [19,20] had an RMSD of 1.095 compared to the template; this estimate takes into account the Cα from the 65 superimposed resi- dues (Figure 6). The global stereochemical quality [21] of the HsZNF706 protein model indicated that 98.3% of the amino acids were in the most allowed and add- itional allowed regions, with only 1.7% (Lys55) in the disallowed region. Structural analysis showed that the side chain of Lys55 is in contact with the solvent in a superficial conformation. The Procheck software [21] was used to calculate the G-factor average value and set to be -0.1. The Verify-3D [22] software analysis, re- vealed that 32.4% of the amino acids had an average value 3D- 1D score of >0.2, showing some compatibi- lity with the HsZNF706 model. Fifty percent of the amino acids in the template had an average value 3D-1D score of >0.2.
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