Heterologous expression and biochemical characterization of an α1,2-mannosidase encoded by the Candida albicans MNS1 gene

Protein glycosylation pathways, commonly found in fungal pathogens, offer an attractive new area of study for the discovery of antifungal targets. In particular, these post-translational modifications are required for virulence and proper cell wall assembly in Candida albicans, an opportunistic human pathogen. The C. albicans MNS1 gene is predicted to encode a member of the glycosyl hydrolase family 47, with 1,2-mannosidase activity. In order to characterise its activity, we first cloned the C. albicans MNS1 gene into Escherichia coli, then expressed and purified the enzyme. The recombinant Mns1 was capable of converting a Man9GlcNAc2 N-glycan core into Man8GlcNAc2 isomer B, but failed to process a Man5GlcNAc2-Asn N-oligosaccharide. These properties are similar to those displayed by Mns1 purified from C. albicansmembranes and strongly suggest that the enzyme is an ±1,2-mannosidase that is localised to the endoplasmic reticulum and involved in the processing of N-linked mannans. Polyclonal antibodies specifically raised against recombinant Mns1 also immunoreacted with the soluble ±1,2-mannosidases E-I and E-II, indicating that Mns1 could share structural similarities with both soluble enzymes. Due to the high degree of similarity between the members of family 47, it is conceivable that these antibodies may recognise ±1,2-mannosidases in other biological systems as well.

Prevalence of antiretroviral drug resistance among treatment-naive and treated HIV-infected patients in Venezuela

An in-house, low-cost method was developed to determine the genotypic resistance of immunodeficiency virus type 1 (HIV-1) isolates. All 179 Venezuelan isolates analysed belonged to subtype B. Primary drug resistance mutations were found in 11% of 63 treatment-naïve patients. The prevalence of resistance in isolates from 116 HIV-positive patients under antiretroviral treatment was 47% to protease inhibitors, 65% to nucleoside inhibitors and 38% to non-nucleoside inhibitors, respectively. Around 50% of patients in the study harboured viruses with highly reduced susceptibility to the three classical types of drugs after only five years from their initial diagnoses.

Purification and biochemica characterisation of endoplasmic reticulum α 1,2-mannosidase from Sporothrix schenckiil

Alpha 1,2-mannosidases from glycosyl hydrolase family 47 participate in N-glycan biosynthesis. In filamentous fungi and mammalian cells, α1,2-mannosidases are present in the endoplasmic reticulum (ER) and Golgi complex and are required to generate complex N-glycans. However, lower eukaryotes such Saccharomyces cerevisiae contain only one α1,2-mannosidase in the lumen of the ER and synthesise high-mannose N-glycans. Little is known about the N-glycan structure and the enzyme machinery involved in the synthesis of these oligosaccharides in the dimorphic fungus Sporothrix schenckii. Here, a membrane-bound α-mannosidase from S. schenckii was solubilised using a high-temperature procedure and purified by conventional methods of protein isolation. Analytical zymograms revealed a polypeptide of 75 kDa to be responsible for enzyme activity and this purified protein was recognised by anti-α1,2-mannosidase antibodies. The enzyme hydrolysed Man9GlcNAc2 into Man8GlcNAc2 isomer B and was inhibited preferentially by 1-deoxymannojirimycin. This α1,2-mannosidase was localised in the ER, with the catalytic domain within the lumen of this compartment. These properties are consistent with an ER-localised α1,2-mannosidase of glycosyl hydrolase family 47. Our results also suggested that in contrast to other filamentous fungi, S. schenckii lacks Golgi α1,2-mannosidases and therefore, the processing of N-glycans by α1,2-mannosidases is similar to that present in lower eukaryotes.

2D-immunoblotting analysis of Sporothrix schenckii cell wall

We utilized two-dimensional gel electrophoresis and immunoblotting (2D-immunoblotting) with anti-Sporothrix schenckii antibodies to identify antigenic proteins in cell wall preparations obtained from the mycelial and yeast-like morphologies of the fungus. Results showed that a 70-kDa glycoprotein (Gp70) was the major antigen detected in the cell wall of both morphologies and that a 60-kDa glycoprotein was present only in yeast-like cells. In addition to the Gp70, the wall from filament cells showed four proteins with molecular weights of 48, 55, 66 and 67 kDa, some of which exhibited several isoforms. To our knowledge, this is the first 2D-immunoblotting analysis of the S. schenckii cell wall.

Insulin receptor substrate-1 expression is increased in circulating leukocytes of patients with acute coronary syndrome.

The mechanisms underlying the increased risk of cardiovascular disease associated with diabetes mellitus (DM) are not fully defined. Insulin resistance in human metabolic syndrome patients is associated with decreased expression of the insulin receptor substrate-2- (Irs2-) AKT2 axis in mononuclear leukocytes (MLs). Moreover, acute coronary syndrome (ACS) has been linked through genome-wide association studies to the 2q36-q37.3 locus, which contains the Irs1 gene. Here, we investigated the expression of insulin-signaling pathway genes in MLs from patients with DM, ACS, and ACS plus DM. Quantitative real-time PCR expression studies showed no differences in the mRNA levels of Irs2, Akt2, and Akt1 among all patients. However, Irs1 mRNA expression was significantly increased in patients with ACS-diabetics and nondiabetics-compared with diabetic patients without ACS (P < .02 and P < .005, resp.). The present study reveals for the first time an association between increased Irs1 mRNA levels in MLs of patients with ACS which is not related to DM.

5-11-19, M. Clemenceau dans le port de Kehl : [photographie de presse] / [Agence Rol]

Référence bibliographique : Rol, 56814

5-11-19, M. Clemenceau arrivant au fort de Kirchbach : [photographie de presse] / [Agence Rol]

Référence bibliographique : Rol, 56810

5-11-19, [M. Clemenceau] dans le port de Kehl : [photographie de presse] / [Agence Rol]

Référence bibliographique : Rol, 56813

Genetic variation in the Cytbgene of human cerebral Taenia soliumcysticerci recovered from clinically and radiologically heterogeneous patients with neurocysticercosis

Neurocysticercosis (NC) is a clinically and radiologically heterogeneous parasitic disease caused by the establishment of larval Taenia solium in the human central nervous system. Host and/or parasite variations may be related to this observed heterogeneity. Genetic differences between pig and human-derived T. solium cysticerci have been reported previously. In this study, 28 cysticerci were surgically removed from 12 human NC patients, the mitochondrial gene that encodes cytochrome b was amplified from the cysticerci and genetic variations that may be related to NC heterogeneity were characterised. Nine different haplotypes (Ht), which were clustered in four haplogroups (Hg), were identified. Hg 3 and 4 exhibited a tendency to associate with age and gender, respectively. However, no significant associations were found between NC heterogeneity and the different T. solium cysticerci Ht or Hg. Parasite variants obtained from patients with similar NC clinical or radiological features were genetically closer than those found in groups of patients with a different NC profile when using the Mantel test. Overall, this study establishes the presence of genetic differences in the Cytb gene of T. solium isolated from human cysticerci and suggests that parasite variation could contribute to NC heterogeneity.

2/6/19, remise du traité de paix aux Autrichiens [préliminaires au traité de Saint-Germain-en-Laye du 10 septembre, au château], maréchal Foch : [photographie de presse] / [Agence Rol]

Référence bibliographique : Rol, 54218

Gravació amb transcodificació al vol des de càmera IP

Desenvolupament d'una aplicació pensada per ser accedida en forma d'aplicatiu web que permeti gestionar gravacions de llarga durada d'imatges i so provinents d'una càmera, fent servir el còdec h.264 per al vídeo i emmagatzemant-lo en un contenidor mp4.