79 research outputs found
Shedding X-ray Light on the Role of Magnesium in the Activity of Mycobacterium tuberculosis Salicylate Synthase (MbtI) for Drug Design
The Mg2+-dependent Mycobacterium tuberculosis salicylate synthase (MbtI) is a key enzyme involved in the biosynthesis of siderophores. Because iron is essential for the survival and pathogenicity of the microorganism, this protein constitutes an attractive target for antitubercular therapy, also considering the absence of homologous enzymes in mammals. An extension of the structure-activity relationships of our furan-based candidates allowed us to disclose the most potent competitive inhibitor known to date (10, Ki = 4 ÎŒM), which also proved effective on mycobacterial cultures. By structural studies, we characterized its unexpected Mg2+-independent binding mode. We also investigated the role of the Mg2+ cofactor in catalysis, analyzing the first crystal structure of the MbtI-Mg2+-salicylate ternary complex. Overall, these results pave the way for the development of novel antituberculars through the rational design of improved MbtI inhibitors
CNTN6 mutations are risk factors for abnormal auditory sensory perception in autism spectrum disorders
Contactin genes CNTN5 and CNTN6 code for neuronal cell adhesion molecules that promote neurite outgrowth in sensory-motor neuronal pathways. Mutations of CNTN5 and CNTN6 have previously been reported in individuals with autism spectrum disorders (ASDs), but very little is known on their prevalence and clinical impact. In this study, we identified CNTN5 and CNTN6 deleterious variants in individuals with ASD. Among the carriers, a girl with ASD and attention-deficit/hyperactivity disorder was carrying five copies of CNTN5. For CNTN6, both deletions (6/1534 ASD vs 1/8936 controls; P=0.00006) and private coding sequence variants (18/501 ASD vs 535/33480 controls; P=0.0005) were enriched in individuals with ASD. Among the rare CNTN6 variants, two deletions were transmitted by fathers diagnosed with ASD, one stop mutation CNTN6W923X was transmitted by a mother to her two sons with ASD and one variant CNTN6P770L was found de novo in a boy with ASD. Clinical investigations of the patients carrying CNTN5 or CNTN6 variants showed that they were hypersensitive to sounds (a condition called hyperacusis) and displayed changes in wave latency within the auditory pathway. These results reinforce the hypothesis of abnormal neuronal connectivity in the pathophysiology of ASD and shed new light on the genes that increase risk for abnormal sensory perception in ASD
Biological and structural characterization of theMycobacterium smegmatis nitroreductase NfnB, and its rolein benzothiazinone resistance
Tuberculosis is still a leading cause of death in developing
countries, for which there is an urgent need
for new pharmacological agents. The synthesis of
the novel antimycobacterial drug class of benzothiazinones
(BTZs) and the identification of their
cellular target as DprE1 (Rv3790), a component of
the decaprenylphosphoryl-b-D-ribose 2'-epimerase
complex, have been reported recently. Here, we
describe the identification and characterization of a
novel resistance mechanism to BTZ in Mycobacterium
smegmatis. The overexpression of the nitroreductase
NfnB leads to the inactivation of the drug by
reduction of a critical nitro-group to an amino-group.
The direct involvement of NfnB in the inactivation of
the lead compound BTZ043 was demonstrated by
enzymology, microbiological assays and gene knockout
experiments. We also report the crystal structure
of NfnB in complex with the essential cofactor flavin
mononucleotide, and show that a common amino
acid stretch between NfnB and DprE1 is likely to be
essential for the interaction with BTZ. We performed
docking analysis of NfnB-BTZ in order to understand
their interaction and the mechanism of
nitroreduction. Although Mycobacterium tuberculosis
seems to lack nitroreductases able to inactivate
these drugs, our findings are valuable for the design
of new BTZ molecules, which may be more effective
in vivo
The bacterial stressosome:a modular system that has been adapted to control secondary messenger signaling
SummaryThe stressosome complex regulates downstream effectors in response to environmental signals. In Bacillus subtilis, it activates the alternative sigma factor ÏB, leading to the upregulation of the general stress regulon. Herein, we characterize a stressosome-regulated biochemical pathway in Moorella thermoacetica. We show that the presumed sensor, MtR, and the scaffold, MtS, form a pseudo-icosahedral structure like that observed in B. subtilis. The N-terminal domain of MtR is structurally homologous to B. subtilis RsbR, despite low sequence identity. The affinity of the switch kinase, MtT, for MtS decreases following MtS phosphorylation and not because of structural reorganization. Dephosphorylation of MtS by the PP2C type phosphatase MtX permits the switch kinase to rebind the stressosome to reset the response. We also show that MtT regulates cyclic di-GMP biosynthesis through inhibition of a GG(D/E)EF-type diguanylate cyclase, demonstrating that secondary messenger levels are regulated by the stressosome
Rotavirus Rearranged Genomic RNA Segments Are Preferentially Packaged into Viruses Despite Not Conferring Selective Growth Advantage to Viruses
The rotavirus (RV) genome consists of 11 double-stranded RNA segments. Sometimes, partial sequence duplication of an RNA segment leads to a rearranged RNA segment. To specify the impact of rearrangement, the replication efficiencies of human RV with rearranged segments 7, 11 or both were compared to these of the homologous human wild-type RV (wt-RV) and of the bovine wt-RV strain RF. As judged by viral growth curves, rotaviruses with a rearranged genome (r-RV) had no selective growth advantage over the homologous wt-RV. In contrast, r-RV were selected over wt-RV during competitive experiments (i.e mixed infections between r-RV and wt-RV followed by serial passages in cell culture). Moreover, when competitive experiments were performed between a human r-RV and the bovine wt-RV strain RF, which had a clear growth advantage, rearranged segments 7, 11 or both always segregated in viral progenies even when performing mixed infections at an MOI ratio of 1 r-RV to 100 wt-RV. Lastly, bovine reassortant viruses that had inherited a rearranged segment 7 from human r-RV were generated. Although substitution of wt by rearranged segment 7 did not result in any growth advantage, the rearranged segment was selected in the viral progenies resulting from mixed infections by bovine reassortant r-RV and wt-RV, even for an MOI ratio of 1 r-RV to 107 wt-RV. Lack of selective growth advantage of r-RV over wt-RV in cell culture suggests a mechanism of preferential packaging of the rearranged segments over their standard counterparts in the viral progeny
Which electrospray-based ionization method best reflects protein-ligand interactions found in solution? A comparison of ESI, nanoESI, and ESSI for the determination of dissociation constants with mass spectrometry
RotavĂrus bovino: fatores de risco, prevalĂȘncia e caracterização antigĂȘnica de amostras em rebanhos leiteiros no estado de SĂŁo Paulo
High prevalence of a variant of SENV in intravenous drug user HIV-infected patients
The prevalence, route of transmission, and clinical significance of SEN virus (SENV) infection was evaluated in 715 samples obtained from 150 blood donors, 165 patients infected by HIV, 150 with HCV/HBV infection, 80 with autoimmune diseases, 40 with Primary Immunodeficiency, 40 with sexually transmitted diseases, 40 polytransfused, and from 50 unselected patients. The identification of SENV-DNA was performed by polymerase chain reaction and hybridization, followed by an immunoenzymatic method that identify different SENV strains. SENV-A variant is largely represented among HIV-infected patients, being found in 71% of HIV+ intravenous drug users and in 26% of individuals that had acquired HIV through sex. A high prevalence of SENV-A was observed also in HIV-polytransfused (27%) or in patients with sexually transmitted diseases (30%). These percentages are significantly higher than those observed in an unselected population and in blood donors. Prevalence of SENV-A is, therefore, high among HIV+ patients with parental risk of exposure, but this infection does not appear to influence the clinical or immune status of HIV+ patients. © 2002 Wiley-Liss, Inc
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