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1.
Int J Mol Sci ; 24(20)2023 Oct 10.
Artículo en Inglés | MEDLINE | ID: mdl-37894729

RESUMEN

Misuse and abuse of antibiotics on humans, cattle, and crops have led to the selection of multi-resistant pathogenic bacteria, the most feared 'superbugs'. Infections caused by superbugs are progressively difficult to treat, with a subsequent increase in lethality: the toll on human lives is predicted to reach 10 million by 2050. Here we review three concepts linked to the growing resistance to antibiotics, namely (i) the Resistome, which refers to the collection of bacterial genes that confer resistance to antibiotics, (ii) the Mobilome, which includes all the mobile genetic elements that participate in the spreading of antibiotic resistance among bacteria by horizontal gene transfer processes, and (iii) the Nichome, which refers to the set of genes that are expressed when bacteria try to colonize new niches. We also discuss the strategies that can be used to tackle bacterial infections and propose an entente cordiale with the bacterial world so that instead of war and destruction of the 'fierce enemy' we can achieve a peaceful coexistence (the One Earth concept) between the human and the bacterial worlds. This, in turn, will contribute to microbial biodiversity, which is crucial in a globally changing climate due to anthropogenic activities.


Asunto(s)
Bacterias , Infecciones Bacterianas , Humanos , Animales , Bovinos , Bacterias/genética , Genes Bacterianos , Infecciones Bacterianas/microbiología , Farmacorresistencia Microbiana , Antibacterianos/farmacología , Farmacorresistencia Bacteriana
2.
Proc Natl Acad Sci U S A ; 114(32): E6526-E6535, 2017 08 08.
Artículo en Inglés | MEDLINE | ID: mdl-28739894

RESUMEN

Relaxases are metal-dependent nucleases that break and join DNA for the initiation and completion of conjugative bacterial gene transfer. Conjugation is the main process through which antibiotic resistance spreads among bacteria, with multidrug-resistant staphylococci and streptococci infections posing major threats to human health. The MOBV family of relaxases accounts for approximately 85% of all relaxases found in Staphylococcus aureus isolates. Here, we present six structures of the MOBV relaxase MobM from the promiscuous plasmid pMV158 in complex with several origin of transfer DNA fragments. A combined structural, biochemical, and computational approach reveals that MobM follows a previously uncharacterized histidine/metal-dependent DNA processing mechanism, which involves the formation of a covalent phosphoramidate histidine-DNA adduct for cell-to-cell transfer. We discuss how the chemical features of the high-energy phosphorus-nitrogen bond shape the dominant position of MOBV histidine relaxases among small promiscuous plasmids and their preference toward Gram-positive bacteria.


Asunto(s)
Proteínas Bacterianas/química , Roturas del ADN de Cadena Simple , ADN Bacteriano/química , Endodesoxirribonucleasas/química , Modelos Moleculares , Plásmidos/química , Staphylococcus aureus/enzimología , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , ADN Bacteriano/genética , ADN Bacteriano/metabolismo , Endodesoxirribonucleasas/genética , Endodesoxirribonucleasas/metabolismo , Histidina/química , Histidina/genética , Histidina/metabolismo , Plásmidos/genética , Plásmidos/metabolismo , Staphylococcus aureus/genética
3.
Nucleic Acids Res ; 45(13): 7774-7785, 2017 Jul 27.
Artículo en Inglés | MEDLINE | ID: mdl-28525572

RESUMEN

Horizontal gene transfer is a key process in the evolution of bacteria and also represents a source of genetic variation in eukaryotes. Among elements participating in gene transfer, thousands of small (<10 kb) mobile bacterial plasmids that replicate by the rolling circle mechanism represent a driving force in the spread of antibiotic resistances. In general, these plasmids are built as genetic modules that encode a replicase, an antibiotic-resistance determinant, and a relaxase that participates in their conjugative mobilization. Further, they control their relatively high copy number (∼30 copies per genome equivalent) by antisense RNAs alone or combined with a repressor protein. We report here that the MobM conjugative relaxase encoded by the promiscuous plasmid pMV158 participates in regulation of the plasmid copy number by transcriptional repression of the antisense RNA, thus increasing the number of plasmid molecules ready to be horizontally transferred (mobilization) and/or vertically inherited (replication). This type of crosstalk between genetic modules involved in vertical and horizontal gene flow has not been reported before.


Asunto(s)
Proteínas Bacterianas/metabolismo , Endodesoxirribonucleasas/metabolismo , Transferencia de Gen Horizontal , Plásmidos/genética , Proteínas Bacterianas/genética , Sitios de Unión , Conjugación Genética , Variaciones en el Número de Copia de ADN , Replicación del ADN , ADN Bacteriano/química , ADN Bacteriano/genética , ADN Bacteriano/metabolismo , ADN Superhelicoidal/química , ADN Superhelicoidal/genética , ADN Superhelicoidal/metabolismo , Farmacorresistencia Bacteriana/genética , Endodesoxirribonucleasas/genética , Escherichia coli/genética , Escherichia coli/metabolismo , Evolución Molecular , Flujo Génico , Microscopía Electrónica , Modelos Biológicos , Regiones Promotoras Genéticas , Replicón , Streptococcus pneumoniae/genética , Streptococcus pneumoniae/metabolismo
4.
Nucleic Acids Res ; 41(14): 6975-91, 2013 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-23723245

RESUMEN

The MgaSpn transcriptional regulator contributes to the virulence of Streptococcus pneumoniae. It is thought to be a member of the Mga/AtxA family of global regulators. MgaSpn was shown to activate in vivo the P1623B promoter, which is divergent from the promoter (Pmga) of its own gene. This activation required a 70-bp region (PB activation region) located between both promoters. In this work, we purified an untagged form of the MgaSpn protein, which formed dimers in solution. By gel retardation and footprinting assays, we analysed the binding of MgaSpn to linear double-stranded DNAs. MgaSpn interacted with the PB activation region when it was placed at internal position on the DNA. However, when it was positioned at one DNA end, MgaSpn recognized preferentially the Pmga promoter placed at internal position. In both cases, and on binding to the primary site, MgaSpn spread along the adjacent DNA regions generating multimeric protein-DNA complexes. When both MgaSpn-binding sites were located at internal positions on longer DNAs, electron microscopy experiments demonstrated that the PB activation region was the preferred target. DNA molecules totally or partially covered by MgaSpn were also visualized. Our results suggest that MgaSpn might recognize particular DNA conformations to achieve DNA-binding specificity.


Asunto(s)
Proteínas Bacterianas/metabolismo , ADN Bacteriano/metabolismo , Streptococcus pneumoniae/genética , Transactivadores/metabolismo , Factores de Virulencia/metabolismo , Proteínas Bacterianas/química , Sitios de Unión , ADN Bacteriano/química , Conformación de Ácido Nucleico , Regiones Promotoras Genéticas , Unión Proteica , Multimerización de Proteína , Streptococcus pneumoniae/patogenicidad , Transactivadores/química , Factores de Virulencia/química
5.
Plasmid ; 74: 15-31, 2014 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-24942190

RESUMEN

Rolling circle-replicating plasmids constitute a vast family that is particularly abundant in, but not exclusive of, Gram-positive bacteria. These plasmids are constructed as cassettes that harbor genes involved in replication and its control, mobilization, resistance determinants and one or two origins of lagging strand synthesis. Any given plasmid may contain all, some, or just only the replication cassette. We discuss here the family of the promiscuous streptococcal plasmid pMV158, with emphasis on its mobilization functions: the product of the mobM gene, prototype of the MOBV relaxase family, and its cognate origin of transfer, oriT. Amongst the subfamily of MOBV1 plasmids, three groups of oriT sequences, represented by plasmids pMV158, pT181, and p1414 were identified. In the same subfamily, we found four types of single-strand origins, namely ssoA, ssoU, ssoW, and ssoT. We found that plasmids of the rolling-circle Rep_2 family (to which pMV158 belongs) are more frequently found in Lactobacillales than in any other bacterial order, whereas Rep_1 initiators seemed to prefer hosts included in the Bacillales order. In parallel, MOBV1 relaxases associated with Rep_2 initiators tended to cluster separately from those linked to Rep_1 plasmids. The updated inventory of MOBV1 plasmids still contains exclusively mobilizable elements, since no genes associated with conjugative transfer (other than the relaxase) were detected. These plasmids proved to have a great plasticity at using a wide variety of conjugative apparatuses. The promiscuous recognition of non-cognate oriT sequences and the role of replication origins for lagging-strand origin in the host range of these plasmids are also discussed.


Asunto(s)
Replicación del ADN/genética , Evolución Molecular , Plásmidos/genética , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Secuencia de Bases , ADN-Topoisomerasas/genética , ADN-Topoisomerasas/metabolismo , ADN Bacteriano/genética , ADN Bacteriano/aislamiento & purificación , Endonucleasas/genética , Endonucleasas/metabolismo , Lactobacillales/genética , Datos de Secuencia Molecular , Conformación de Ácido Nucleico , Origen de Réplica/genética , Streptococcus/genética
6.
Int J Vitam Nutr Res ; 84(1-2): 92-7, 2014.
Artículo en Inglés | MEDLINE | ID: mdl-25835239

RESUMEN

INTRODUCTION: Low maternal vitamin B12 status is a risk factor for various adverse pregnancy outcomes. Although vitamin B12 deficiency is not a primary target of newborn screening (NBS) programs, measurements of propionylcarnitine (C3) and its ratios with acetylcarnitine (C3/C2) and palmitoylcarnitine (C3/C16) may incidentally identify vitamin B12-deficient newborns. The objective of this study was to measure vitamin B12 levels in women during the first trimester of pregnancy, evaluate predictors of these concentrations, and study their relationship with newborn screening results. DESIGN: Vitamin B12 concentrations were evaluated in 204 women during the first trimester of pregnancy and possible confounding factors were analyzed. After giving birth, data of their newborns (189) were collected (sex, gestational age, birthweight) and the acylcarnitine profile obtained by tandem mass spectrometry during NBS was analyzed. To assess the effects of the variables on vitamin B12 serum concentrations and newborn screening markers, stepwise multiple linear regression models were used. RESULTS: The mean serum concentration of vitamin B12 was 370.8 pmol/L (502.4 pg/mL) (SD 142.81). Vitamin B12 concentrations were significantly lower in smokers (p=0.027), and in women with low meat consumption (p=0.040). There was a significant inverse correlation between mothers'’ vitamin B12 concentrations and their children’'s C3 (r=-0.24; p=0.001), C3/C2 (r=-0.23; p=0.002) and C3/C16 levels (r=-0.20; p=0.006). CONCLUSIONS: Newborn screening markers (C3, C3/C2, and C3/C16) present an inverse correlation with maternal vitamin B12 status in the first trimester of pregnancy. Regarding factors that may influence maternal serum vitamin B12 levels during the first trimester, smoking seems to have a negative effect, and meat consumption a positive effect.


Asunto(s)
Biomarcadores/sangre , Tamizaje Neonatal , Deficiencia de Vitamina B 12/sangre , Vitamina B 12/sangre , Acetilcarnitina/sangre , Adolescente , Adulto , Carnitina/análogos & derivados , Carnitina/sangre , Dieta , Femenino , Edad Gestacional , Humanos , Recién Nacido , Bienestar Materno , Carne , Estado Nutricional , Embarazo , Complicaciones del Embarazo/sangre , Tercer Trimestre del Embarazo , Fumar/efectos adversos , Fumar/sangre , Adulto Joven
7.
Parasit Vectors ; 17(1): 331, 2024 Aug 06.
Artículo en Inglés | MEDLINE | ID: mdl-39107844

RESUMEN

BACKGROUND: Aedes aegypti (L.) is the main vector of dengue, yellow fever, Zika, and chikungunya viruses in many parts of the world, impacting millions of people worldwide each year. Insecticide-based interventions have been effective in controlling Aedes mosquito populations for several years, but in recent times, resistance to these compounds has developed, posing a global threat to the control of this mosquito. METHODS: Ovitraps were used to collect A. aegypti eggs in the cities of Tartagal and San Ramón de la Nueva Orán (Salta), Puerto Iguazú (Misiones), and Clorinda (Formosa). World Health Organization (WHO)-impregnated papers with the discriminating concentration (DC) of permethrin, 5X, 10X and pirimiphos methyl were used for the toxicological bioassays. We also genotyped each sample for the three kdr single nucleotide polymorphisms (SNP): V410L, V1016I, and F1534C in individual TaqMan quantitative PCR (qPCR) reactions. RESULTS: All investigated A. aegypti populations were highly resistant to permethrin, as the mortality percentage with the permethrin 10×DC remained below 98%. However, all populations were 100% susceptible to pirimiphos-methyl. Kdr genotyping demonstrated the presence of the V410L mutation for the first time in Argentina in all the populations studied. A prevalence of the triple mutant genotype (LL + II + CC) was observed in the northeastern cities of Clorinda (83.3%) and Puerto Iguazú (55.6%). CONCLUSIONS: This study demonstrates for the first time the presence and intensity of resistance to permethrin in different populations from Argentina, and correlates the observed phenotype with the presence of kdr mutations (genotype).


Asunto(s)
Aedes , Resistencia a los Insecticidas , Insecticidas , Mosquitos Vectores , Mutación , Aedes/efectos de los fármacos , Aedes/genética , Animales , Argentina , Resistencia a los Insecticidas/genética , Insecticidas/farmacología , Mosquitos Vectores/genética , Mosquitos Vectores/efectos de los fármacos , Permetrina/farmacología , Polimorfismo de Nucleótido Simple , Genotipo
8.
J Bacteriol ; 195(13): 3000-8, 2013 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-23625844

RESUMEN

A crucial element in the horizontal transfer of mobilizable and conjugative plasmids is the relaxase, a single-stranded endonuclease that nicks the origin of transfer (oriT) of the plasmid DNA. The relaxase of the pMV158 mobilizable plasmid is MobM (494 residues). In solution, MobM forms a dimer through its C-terminal domain, which is proposed to anchor the protein to the cell membrane and to participate in type 4 secretion system (T4SS) protein-protein interactions. In order to gain a deeper insight into the structural MobM requirements for efficient DNA catalysis, we studied two endonuclease domain variants that include the first 199 or 243 amino acid residues (MobMN199 and MobMN243, respectively). Our results confirmed that the two proteins behaved as monomers in solution. Interestingly, MobMN243 relaxed supercoiled DNA and cleaved single-stranded oligonucleotides harboring oriTpMV158, whereas MobMN199 was active only on supercoiled DNA. Protein stability studies using gel electrophoresis and mass spectrometry showed increased susceptibility to degradation at the domain boundary between the N- and C-terminal domains, suggesting that the domains change their relative orientation upon DNA binding. Overall, these results demonstrate that MobMN243 is capable of nicking the DNA substrate independently of its topology and that the amino acids 200 to 243 modulate substrate specificity but not the nicking activity per se. These findings suggest that these amino acids are involved in positioning the DNA for the nuclease reaction rather than in the nicking mechanism itself.


Asunto(s)
Proteínas Bacterianas/química , Proteínas Bacterianas/metabolismo , Endodesoxirribonucleasas/química , Endodesoxirribonucleasas/metabolismo , Streptococcus pneumoniae/metabolismo , Proteínas Bacterianas/genética , ADN/metabolismo , ADN Superhelicoidal/metabolismo , Endodesoxirribonucleasas/genética , Plásmidos/genética , Unión Proteica
9.
EMBO J ; 28(11): 1666-78, 2009 Jun 03.
Artículo en Inglés | MEDLINE | ID: mdl-19440202

RESUMEN

RepB initiates plasmid rolling-circle replication by binding to a triple 11-bp direct repeat (bind locus) and cleaving the DNA at a specific distant site located in a hairpin loop within the nic locus of the origin. The structure of native full-length RepB reveals a hexameric ring molecule, where each protomer has two domains. The origin-binding and catalytic domains show a three-layer alpha-beta-alpha sandwich fold. The active site is positioned at one of the faces of the beta-sheet and coordinates a Mn2+ ion at short distance from the essential nucleophilic Y99. The oligomerization domains (ODs), each consisting of four alpha-helices, together define a compact ring with a central channel, a feature found in ring helicases. The toroidal arrangement of RepB suggests that, similar to ring helicases, it encircles one of the DNA strands during replication to confer processivity to the replisome complex. The catalytic domains appear to be highly mobile with respect to ODs. This mobility may account for the adaptation of the protein to two distinct DNA recognition sites.


Asunto(s)
ADN Helicasas/química , ADN Helicasas/metabolismo , Subunidades de Proteína/química , Secuencia de Aminoácidos , Sitios de Unión , Dominio Catalítico , Modelos Moleculares , Datos de Secuencia Molecular , Multimerización de Proteína , Estructura Cuaternaria de Proteína , Estructura Secundaria de Proteína , Estructura Terciaria de Proteína
10.
Plasmid ; 70(1): 120-30, 2013 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-23562993

RESUMEN

The MobM relaxase (494 amino acids) encoded by the promiscuous streptococcal plasmid pMV158 recognizes the plasmid origin of transfer, oriTpMV158, and converts supercoiled pMV158 DNA into relaxed molecules by cleavage of the phosphodiester bond of a specific dinucleotide within the sequence 5'-GTGTG/TT-3' ("/" being the nick site). After cleavage, the protein remains stably bound to the 5'-end of the nick site. Band-shift assays with single-stranded oligonucleotides and size-exclusion chromatography allowed us to show that MobM was able to generate specific complexes with one of the inverted repeats of the oriTpMV158, presumably extruded as stem-loop structure. A number of tests have been performed to attain a better characterization of the nicking activity of MobM and its linkage with its target DNA. The optimal pH for DNA relaxation was found to be 6.5. Upon nicking, gel retardation assays showed that MobM formed stable complexes with its target DNA. Moreover, MobM bound to relaxed pMV158 molecules were visualized by electron microscopy. The staphylococcal plasmids pUB110 and pE194, and the streptococcal plasmid pDL287 harbour putative oriTs and may encode Mob proteins homologous to MobM. The oriTpUB110, oriTpDL287, and oriTpE194 sequences share 100%, 70%, and 67% (in a 43-nucleotide stretch and allowing a 3-bp gap) identity to oriTpMV158, respectively. Nicking assays using supercoiled DNAs from pUB110, pDL287, and pE194 showed that MobM was able to relax, to differing degrees, all plasmid DNAs. Our results suggest that cross-recognition of heterologous oriTs by Mob proteins could play an important role in the plasmid spreading between bacteria.


Asunto(s)
Proteínas Bacterianas/genética , ADN Bacteriano/genética , ADN Superhelicoidal/genética , Endodesoxirribonucleasas/genética , Plásmidos/genética , Streptococcus pneumoniae/genética , Secuencia de Aminoácidos , Proteínas Bacterianas/metabolismo , Sitios de Unión , Conjugación Genética , ADN Bacteriano/química , ADN Bacteriano/metabolismo , ADN Superhelicoidal/química , ADN Superhelicoidal/metabolismo , Ensayo de Cambio de Movilidad Electroforética , Endodesoxirribonucleasas/metabolismo , Concentración de Iones de Hidrógeno , Secuencias Invertidas Repetidas , Datos de Secuencia Molecular , Conformación de Ácido Nucleico , Plásmidos/química , Plásmidos/metabolismo , Unión Proteica , Homología de Secuencia de Ácido Nucleico , Streptococcus pneumoniae/enzimología
11.
Nucleic Acids Res ; 39(10): 4315-29, 2011 May.
Artículo en Inglés | MEDLINE | ID: mdl-21296755

RESUMEN

Protein MobM, the relaxase involved in conjugative transfer of the streptococcal plasmid pMV158, is the prototype of the MOB(V) superfamily of relaxases. To characterize the DNA-binding and nicking domain of MobM, a truncated version of the protein (MobMN199) encompassing its N-terminal region was designed and the protein was purified. MobMN199 was monomeric in contrast to the dimeric form of the full-length protein, but it kept its nicking activity on pMV158 DNA. The optimal relaxase activity was dependent on Mn(2+) or Mg(2+) cations in a dosage-dependent manner. However, whereas Mn(2+) strongly stabilized MobMN199 against thermal denaturation, no protective effect was observed for Mg(2+). Furthermore, MobMN199 exhibited a high affinity binding for Mn(2+) but not for Mg(2+). We also examined the binding-specificity and affinity of MobMN199 for several substrates of single-stranded DNA encompassing the pMV158 origin of transfer (oriT). The minimal oriT was delimited to a stretch of 26 nt which included an inverted repeat located eight bases upstream of the nick site. The structure of MobMN199 was strongly stabilized by binding to the defined target DNA, indicating the formation of a tight protein-DNA complex. We demonstrate that the oriT recognition by MobMN199 was highly specific and suggest that this protein most probably employs Mn(2+) during pMV158 transfer.


Asunto(s)
Proteínas Bacterianas/química , ADN de Cadena Simple/metabolismo , Endodesoxirribonucleasas/química , Manganeso/química , Plásmidos/genética , Secuencia de Aminoácidos , Proteínas Bacterianas/metabolismo , Cationes/química , Endodesoxirribonucleasas/metabolismo , Estabilidad de Enzimas , Manganeso/farmacología , Datos de Secuencia Molecular , Plásmidos/metabolismo , Estructura Secundaria de Proteína , Estructura Terciaria de Proteína , Temperatura
12.
FEMS Microbiol Rev ; 47(5)2023 09 05.
Artículo en Inglés | MEDLINE | ID: mdl-37715317

RESUMEN

Toxin-antitoxin (TA) systems are entities found in the prokaryotic genomes, with eight reported types. Type II, the best characterized, is comprised of two genes organized as an operon. Whereas toxins impair growth, the cognate antitoxin neutralizes its activity. TAs appeared to be involved in plasmid maintenance, persistence, virulence, and defence against bacteriophages. Most Type II toxins target the bacterial translational machinery. They seem to be antecessors of Higher Eukaryotes and Prokaryotes Nucleotide-binding (HEPN) RNases, minimal nucleotidyltransferase domains, or CRISPR-Cas systems. A total of four TAs encoded by Streptococcus pneumoniae, RelBE, YefMYoeB, Phd-Doc, and HicAB, belong to HEPN-RNases. The fifth is represented by PezAT/Epsilon-Zeta. PezT/Zeta toxins phosphorylate the peptidoglycan precursors, thereby blocking cell wall synthesis. We explore the body of knowledge (facts) and hypotheses procured for Type II TAs and analyse the data accumulated on the PezAT family. Bioinformatics analyses showed that homologues of PezT/Zeta toxin are abundantly distributed among 14 bacterial phyla mostly in Proteobacteria (48%), Firmicutes (27%), and Actinobacteria (18%), showing the widespread distribution of this TA. The pezAT locus was found to be mainly chromosomally encoded whereas its homologue, the tripartite omega-epsilon-zeta locus, was found mostly on plasmids. We found several orphan pezT/zeta toxins, unaccompanied by a cognate antitoxin.


Asunto(s)
Antitoxinas , Toxinas Bacterianas , Antitoxinas/química , Antitoxinas/genética , Toxinas Bacterianas/genética , Bacterias/genética , Bacterias/metabolismo , Operón , Células Procariotas/metabolismo , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo
13.
Front Mol Biosci ; 10: 1294974, 2023.
Artículo en Inglés | MEDLINE | ID: mdl-38192335

RESUMEN

When Enterococcus faecalis is exposed to changing environmental conditions, the expression of many genes is regulated at the transcriptional level. We reported previously that the enterococcal MafR protein causes genome-wide changes in the transcriptome. Here we show that MafR activates directly the transcription of the OG1RF_10478 gene, which encodes a hypothetical protein of 111 amino acid residues. We have identified the P10478 promoter and demonstrated that MafR enhances the efficiency of this promoter by binding to a DNA site that contains the -35 element. Moreover, our analysis of the OG1RF_10478 protein AlphaFold model indicates high similarity to 1) structures of EIIB components of the bacterial phosphoenolpyruvate:carbohydrate phosphotransferase system, and 2) structures of receiver domains that are found in response regulators of two-component signal transduction systems. However, unlike typical EIIB components, OG1RF_10478 lacks a Cys or His residue at the conserved phosphorylation site, and, unlike typical receiver domains, OG1RF_10478 lacks a conserved Asp residue at the position usually required for phosphorylation. Different from EIIB components and receiver domains, OG1RF_10478 contains an insertion between residues 10 and 30 that, according to ColabFold prediction, may serve as a dimerization interface. We propose that OG1RF_10478 could participate in regulatory functions by protein-protein interactions.

14.
Trop Med Infect Dis ; 8(7)2023 Jul 14.
Artículo en Inglés | MEDLINE | ID: mdl-37505658

RESUMEN

Strategies for the prevention of arboviral diseases transmitted by Aedes aegypti have traditionally focused on vector control. This remains the same to this day, despite a lack of documented evidence on its efficacy due to a lack of coverage and sustainability. The continuous growth of urban areas and generally unplanned urbanization, which favor the presence of Ae. aegypti, demand resources, both material and human, as well as logistics to effectively lower the population's risk of infection. These considerations have motivated the development of tools to identify areas with a recurrent concentration of arboviral cases during an outbreak to be able to prioritize preventive actions and optimize available resources. This study explores the existence of spatial patterns of dengue incidence in the locality of Tartagal, in northeastern Argentina, during the outbreaks that occurred between 2010 and 2020. Approximately half (50.8%) of the cases recorded during this period were concentrated in 35.9% of the urban area. Additionally, an important overlap was found between hotspot areas of dengue and chikungunya (Kendall's W = 0.92; p-value < 0.001) during the 2016 outbreak. Moreover, 65.9% of the cases recorded in 2022 were geolocalized within the hotspot areas detected between 2010 and 2020. These results can be used to generate a risk map to implement timely preventive control strategies that prioritize these areas to reduce their vulnerability while optimizing the available resources and increasing the scope of action.

15.
J Bacteriol ; 194(16): 4197-207, 2012 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-22661692

RESUMEN

Global transcriptional regulators that respond to specific environmental signals are crucial in bacterial pathogenesis. In the case of the Gram-positive pathogen Streptococcus pneumoniae (the pneumococcus), the sp1800 gene of the clinical isolate TIGR4 encodes a protein that exhibits homology to the Mga "stand-alone" response regulator of the group A Streptococcus. Such a pneumococcal protein was shown to play a significant role in both nasopharyngeal colonization and development of pneumonia in murine infection models. Moreover, it was shown to repress the expression of several genes located within the rlrA pathogenicity islet. The pneumococcal R6 strain, which derives from the D39 clinical isolate, lacks the rlrA islet but has a gene (here named mga(Spn)) equivalent to the sp1800 gene. In this work, and using in vivo approaches, we have identified the promoter of the mga(Spn) gene (Pmga) and demonstrated that four neighboring open reading frames of unknown function (spr1623 to spr1626) constitute an operon. Transcription of this operon is under the control of two promoters (P1623A and P1623B) that are divergent from the Pmga promoter. Furthermore, we have shown that the Mga(Spn) protein activates the P1623B promoter in vivo. This activation requires sequences located around 50 to 120 nucleotides upstream of the P1623B transcription start site. By DNase I footprinting assays, we have also demonstrated that such a region includes an Mga(Spn) binding site. This is the first report on the activator role of the pneumococcal Mga-like protein.


Asunto(s)
Proteínas Bacterianas/metabolismo , Streptococcus pneumoniae/genética , Factores de Transcripción/metabolismo , Activación Transcripcional , Factores de Virulencia/biosíntesis , Huella de ADN , Operón , Regiones Promotoras Genéticas , Streptococcus pneumoniae/metabolismo , Transcripción Genética , Virulencia
16.
J Bacteriol ; 194(7): 1789-99, 2012 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-22287528

RESUMEN

The streptococcal promiscuous plasmid pMV158 (5,540 bp) replicates by the rolling-circle mechanism and can be mobilized among a wide number of Gram-positive and -negative bacteria. The plasmid region involved in its conjugative transfer includes the mobM gene, which encodes the MobM relaxase, and the cis-acting origin of transfer (oriT). MobM initiates transfer by cleavage of supercoiled pMV158 DNA at a specific dinucleotide within oriT. In the present work, we have performed a detailed transcriptional analysis to assess the role of MobM in the control of its own gene expression. By in vivo and in vitro approaches, we demonstrated that mobM transcription in Escherichia coli was mostly initiated from a promoter (Pmob2) different from the one (Pmob1) used in Lactococcus lactis. Whereas promoter Pmob1 was embedded within the oriT sequence, promoter Pmob2 was placed apart from but adjacent to oriT. Further, MobM was able to repress the expression of its own gene from both promoters. Given the promiscuity of pMV158, the organization of the mobM promoter region suggests a strategy of the plasmid to cope with different transcription machineries of the hosts it colonizes.


Asunto(s)
Proteínas Bacterianas/genética , Endodesoxirribonucleasas/genética , Escherichia coli/genética , Regulación Enzimológica de la Expresión Génica , Lactococcus lactis/genética , Plásmidos/genética , Biosíntesis de Proteínas , Proteínas Bacterianas/metabolismo , Conjugación Genética , Endodesoxirribonucleasas/metabolismo , Escherichia coli/enzimología , Regulación Bacteriana de la Expresión Génica , Lactococcus lactis/enzimología , Datos de Secuencia Molecular , Plásmidos/metabolismo , Regiones Promotoras Genéticas , Transcripción Genética
17.
Proteins ; 80(7): 1834-46, 2012 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-22488579

RESUMEN

The chromosome of the pathogenic Gram-positive bacterium Streptococcus pneumoniae contains between six to 10 operons encoding toxin-antitoxin systems (TAS). TAS are widespread and redundant in bacteria and archaea and their role, albeit still obscure, may be related to important aspects of bacteria lifestyle like response to stress. One of the most abundant TAS is the relBE family, being present in the chromosome of many bacteria and archaea. Because of the high rates of morbility and mortality caused by S. pneumoniae, it has been interesting to gain knowledge on the pneumococcal TAS, among them the RelBE2Spn proteins. Here, we have analyzed the DNA binding capacity of the RelB2Spn antitoxin and the RelB2Spn-RelE2Spn proteins by band-shift assays. Thus, a DNA region encompassing the operator region of the proteins was identified. In addition, we have used analytical ultracentrifugation and native mass spectrometry to measure the oligomerization state of the antitoxin alone and the RelBE2Spn complex in solution bound or unbound to its DNA substrate. Using native mass spectrometry allowed us to unambiguously determine the stoichiometry of the RelB2Spn and of the RelBE2Spn complex alone or associated to its DNA target.


Asunto(s)
Proteínas Bacterianas/química , Toxinas Bacterianas/química , Proteínas de Unión al ADN/química , ADN/química , Streptococcus pneumoniae/química , Secuencia de Aminoácidos , Proteínas Bacterianas/metabolismo , Toxinas Bacterianas/metabolismo , Secuencia de Bases , ADN/metabolismo , Proteínas de Unión al ADN/metabolismo , Espectrometría de Masas , Datos de Secuencia Molecular , Complejos Multiproteicos/química , Complejos Multiproteicos/metabolismo , Unión Proteica , Multimerización de Proteína , Proteínas Recombinantes/química , Proteínas Recombinantes/aislamiento & purificación , Proteínas Recombinantes/metabolismo , Ultracentrifugación
18.
Plasmid ; 67(2): 162-6, 2012 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-22252136

RESUMEN

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.


Asunto(s)
Plásmidos/genética , Replicón/genética , Streptococcus pneumoniae/crecimiento & desarrollo , Streptococcus pneumoniae/genética , Orden Génico , Plásmidos/metabolismo
19.
Plasmid ; 67(1): 53-9, 2012 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-21946126

RESUMEN

We report the construction of a plasmid vector designed for regulated gene expression in Streptococcus pneumoniae. The new vector, pLS1ROM, is based on the replicon of the streptococcal promiscuous rolling circle replication (RCR) plasmid pMV158. We inserted the controllable promoter P(M) of the S. pneumoniaemalMP operon, followed by a multi-cloning site sequence aimed to facilitate the insertion of target genes. The expression from P(M) is negatively regulated by the transcriptional repressor MalR, which is released from the DNA operator sequence by growing the cells in maltose-containing media. To get a highly regulated expression of the target gene, MalR was provided in cis by inserting the malR gene under control of the constitutive P(tet) promoter, which in pMV158 directs expression of the tetL gene. To test the functionality of the system, we cloned the reporter gene gfp from Aequorea victoria, encoding the green fluorescent protein (GFP). Pneumococcal cells harboring the recombinant plasmid rendered GFP fluorescence in a maltose-dependent mode with undetectable background levels in the absence of the inducer. The new vector, pLS1ROM, exhibits full structural and segregational stability and constitutes a valuable tool for genetic manipulation and regulated gene expression in S. pneumoniae.


Asunto(s)
Regulación Bacteriana de la Expresión Génica , Genes Bacterianos/genética , Vectores Genéticos , Proteínas Fluorescentes Verdes/genética , Plásmidos/genética , Regiones Promotoras Genéticas/genética , Replicón/genética , Streptococcus pneumoniae/genética , Proteínas Bacterianas/genética , Secuencia de Bases , Clonación Molecular , ADN Bacteriano/genética , Proteínas Fluorescentes Verdes/metabolismo , Maltosa/metabolismo , Datos de Secuencia Molecular , Reacción en Cadena de la Polimerasa , Proteínas Represoras/genética , Homología de Secuencia de Ácido Nucleico
20.
Front Mol Biosci ; 9: 848444, 2022.
Artículo en Inglés | MEDLINE | ID: mdl-35402507

RESUMEN

The biological-biochemical community has been shocked and delighted by the remarkable progress that has recently been made on a problem that has consumed the attention, energy, and resources of many, if not most of the scientists in the field for the past 50 years. The problem has been to predict the tertiary structure of a protein merely from its amino acid sequence. Nature does it easily enough, but it has been an incredibly difficult problem, often considered intractable, for humankind. The breakthrough has come in the form of two computer-based approaches, AlphaFold2 and RoseTTAFold in conjunction with factors such as the use of vast computing power, the field of artificial intelligence, and the existence of huge protein sequence databases. The advancement of these tools depended upon and was stimulated by the last 50 years of development of smaller and smaller and more and more powerful electronics components, mainly processors and memory. Along with the problem of protein folding, determining the function or mechanism of action of proteins has similarly limped along as did protein folding until the recent breakthroughs. Perhaps AlphaFold2 and RoseTTAFold can substantially aid in protein mechanistic studies. Now it is not completely insane to consider what might be the next grand challenge in biochemistry-biology. We offer several possibilities.

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