Role of histidine-50, glutamic acid-96, and histidine-137 in the ribonucleolytic mechanism of the ribotoxin alpha-sarcin

alpha-Sarcin is a ribotoxin secreted by the mold Aspergillus giganteus that degrades the ribosomal RNA by acting as a cyclizing ribonuclease. Three residues potentially involved in the mechanism of catalysis--histidine-50, glutamic acid-96, and histidine-137--were changed to glutamine. Three different single mutation variants (H50Q, E96Q, H137Q) as well as a double variant (H50/137Q) and a triple variant (H50/137Q/E96Q) were prepared and isolated to homogeneity. These variants were spectroscopically (circular dichroism, fluorescence emission, and proton nuclear magnetic resonance) characterized. According to these results, the three-dimensional structure of these variants of alpha-sarcin was preserved; only very minor local changes were detected. All the variants were inactive when assayed against either intact ribosomes or poly(A). The effect of pH on the ribonucleolytic activity of alpha-sarcin was evaluated against the ApA dinucleotide. This assay revealed that only the H50Q variant still retained its ability to cleave a phosphodiester bond, but it did so to a lesser extent than did wild-type alpha-sarcin. The results obtained are interpreted in terms of His137 and Glu96 as essential residues for the catalytic activity of alpha-sarcin (His137 as the general acid and Glu96 as the general base) and His50 stabilizing the transition state of the reaction catalyzed by alpha-sarcin.     

A strictly anaerobic betaproteobacterium Georgfuchsia toluolica gen. nov., sp. nov. degrades aromatic compounds with Fe(III), Mn(IV) or nitrate as an electron acceptor

A bacterium (strain G5G6) that grows anaerobically with toluene was isolated from a polluted aquifer (Banisveld, the Netherlands). The bacterium uses Fe(III), Mn(IV) and nitrate as terminal electron acceptors for growth on aromatic compounds. The bacterium does not grow on sugars, lactate or acetate. Phylogenetic analysis of the 16S rRNA gene sequence indicated that strain G5G6 belonged to the Betaproteobacteria . Its closest, but only distantly related, cultured relative is Sterolibacterium denitrificans Chol-1S^T (94.6% similarity of the 16S rRNA genes), a cholesterol-oxidizing, denitrifying bacterium. Strain G5G6 possesses the benzylsuccinate synthase A ( bssA ) gene encoding the α-subunit of Bss, which catalyzes the first step in anaerobic toluene degradation. The deduced BssA amino acid sequence is closely related to those of Azoarcus and Thauera species, which also belong to the Betaproteobacteria . Strain G5G6 is the first toluene-degrading, iron-reducing bacterium that does not belong to the Geobacteraceae within the Deltaproteobacteria . Based on phylogenetic and physiological comparison, strain G5G6 could not be assigned to a described species. Therefore, strain G5G6 (DSMZ 19032^T=JCM 14632^T) is a novel taxon of the Betaproteobacteria . We propose the name Georgfuchsia toluolica gen. nov., sp. nov.     

The formation of titan cells in Cryptococcus neoformans depends on the mouse strain and correlates with induction of Th2‐type responses

Cryptococcus neoformans is a pathogenic yeast that can form titan cells in the lungs, which are fungal cells of abnormal enlarged size. Little is known about the factors that trigger titan cells. In particular, it is not known how the host environment influences this transition. In this work, we describe the formation of titan cells in two mouse strains, CD1 and C57BL/6J. We found that the proportion of C. neoformans titan cells was significantly higher in C57BL/6J mice than in CD1. This higher proportion of titan cells was associated with a higher dissemination of the yeasts to the brain. Histology sections demonstrated eosinophilia in infected animals, although it was significantly lower in the CD1 mice which presented infiltration of lymphocytes. Both mouse strains presented infiltration of granulocytes, but the amount of eosinophils was higher in C57BL/6J. CD1 mice showed a significant accumulation of IFN‐γ, TNF‐α and IL17, while C57BL/BL mice had an increase in the anti‐inflammatory cytokine IL‐4. IgM antibodies to the polysaccharide capsule and total IgE were more abundant in the sera from C57BL/6J, confirming that these animals present a Th2‐type response. We conclude that titan cell formation in C. neoformans depends, not only on microbe factors, but also on the host environment.     

Multiscale modelling of Potts shunt as a potential palliative treatment for suprasystemic idiopathic pulmonary artery hypertension: a paediatric case study

Biomechanics and Modeling in Mechanobiology - Potts shunt (PS) was suggested as palliation for patients with suprasystemic pulmonary arterial hypertension (PAH) and right ventricular (RV) failure....     

Fitness of the pMV158 replicon in Streptococcus pneumoniae

Promiscuous, rolling-circle replication plasmid pMV158 determines tetracycline resistance to its host and can be mobilized by conjugation. Plasmid pLS1 is a deletion derivative of pMV158 that has lost its conjugative mobilization ability. Both plasmids replicate efficiently and are stably inherited in Streptococcus pneumoniae. We have analyzed the effect of pMV158 and pLS1 carriage on the bacterial growth rate. Whereas the parental plasmid does not significantly modify the cell doubling time, pLS1 slows down the growth of the bacterial host by 8-9%. The bases of the differential burden caused by pMV158 and pLS1 carriage are not yet understood. The negligible cost of the pMV158 parental natural plasmid on the host might explain the prevalence of small, multicopy, rolling-circle replication plasmids, even though they lack any selectable trait.