RESUMEN
BACKGROUND: PAI-1 gain-of-function variants promote airway fibrosis and are associated with asthma and with worse lung function in subjects with asthma. OBJECTIVE: We sought to determine whether the association of a gain-of-function polymorphism in plasminogen activator inhibitor-1 (PAI-1) with airway obstruction is modified by asthma status, and whether any genotype effect persists after accounting for common exposures that increase PAI-1 level. METHODS: We studied 2070 Latino children (8-21y) with genotypic and pulmonary function data from the GALA II cohort. We estimated the relationship of the PAI-1 risk allele with FEV1/FVC by multivariate linear regression, stratified by asthma status. We examined the association of the polymorphism with asthma and airway obstruction within asthmatics via multivariate logistic regression. We replicated associations in the SAPPHIRE cohort of African Americans (n=1056). Secondary analysis included the effect of the at-risk polymorphism on postbronchodilator lung function. RESULTS: There was an interaction between asthma status and the PAI-1 polymorphism on FEV1 /FVC (P=.03). The gain-of-function variants, genotypes (AA/AG), were associated with lower FEV1 /FVC in subjects with asthma (ß=-1.25, CI: -2.14,-0.35, P=.006), but not in controls. Subjects with asthma and the AA/AG genotypes had a 5% decrease in FEV1 /FVC (P<.001). In asthmatics, the risk genotype (AA/AG) was associated with a 39% increase in risk of clinically relevant airway obstruction (OR=1.39, CI: 1.01, 1.92, P=.04). These associations persisted after exclusion of factors that increase PAI-1 including tobacco exposure and obesity. CONCLUSIONS AND CLINICAL RELEVANCE: The decrease in the FEV1 /FVC ratio associated with the risk genotype was modified by asthma status. The genotype increased the odds of airway obstruction by 75% within asthmatics only. As exposures known to increase PAI-1 levels did not mitigate this association, PAI-1 may contribute to airway obstruction in the context of chronic asthmatic airway inflammation.
Asunto(s)
Obstrucción de las Vías Aéreas/genética , Obstrucción de las Vías Aéreas/metabolismo , Mutación con Ganancia de Función , Inhibidor 1 de Activador Plasminogénico/genética , Inhibidor 1 de Activador Plasminogénico/metabolismo , Adolescente , Adulto , Obstrucción de las Vías Aéreas/epidemiología , Obstrucción de las Vías Aéreas/fisiopatología , Alelos , Asma Ocupacional/epidemiología , Asma Ocupacional/genética , Asma Ocupacional/metabolismo , Asma Ocupacional/fisiopatología , Niño , Estudios de Cohortes , Etnicidad , Femenino , Estudios de Asociación Genética , Predisposición Genética a la Enfermedad , Genotipo , Humanos , Masculino , Oportunidad Relativa , Polimorfismo de Nucleótido Simple , Pruebas de Función Respiratoria , Adulto JovenRESUMEN
INTRODUCTION: Mixed dementia (DMix) refers to dementia resulting from Alzheimer disease in addition to cerebrovascular disease. The study objectives were to determine the clinical and imaging factors associated with Dmix and compare them to those associated with Alzheimer disease. MATERIAL AND METHODS: Cross-sectional study including 225 subjects aged 65 years and over from a memory clinic in a tertiary hospital in Mexico City. All patients underwent clinical, neuropsychological, and brain imaging studies. We included patients diagnosed with DMix or Alzheimer disease (AD). A multivariate analysis was used to determine factors associated with DMix. RESULTS: We studied 137 subjects diagnosed with Dmix. Compared to patients with AD, Dmix patients were older and more likely to present diabetes, hypertension, dyslipidaemia, and history of cerebrovascular disease (P<.05). The multivariate analysis showed that hypertension (OR 1.92, CI 1.62-28.82; P=.009), white matter disease (OR 3.61, CI 8.55-159.80; P<.001), and lacunar infarcts (OR 3.35, CI 1.97-412.34; P=.014) were associated with Dmix, whereas a history of successfully treated depression showed an inverse association (OR 0.11, CI 0.02-0-47; P=.004) CONCLUSIONS: DMix may be more frequent than AD. Risk factors such as advanced age and other potentially modifiable factors were associated with this type of dementia. Clinicians should understand and be able to define Dmix.
Asunto(s)
Enfermedad de Alzheimer/complicaciones , Trastornos Cerebrovasculares/complicaciones , Anciano de 80 o más Años , Trastornos Cerebrovasculares/etiología , Estudios Transversales , Demencia Vascular , Femenino , Humanos , Masculino , México , Factores de RiesgoRESUMEN
BACKGROUND: Younger maternal age at birth is associated with increased risk of asthma in offspring in European descent populations, but has not been studied in Latino populations. OBJECTIVES: We sought to examine the relationship between maternal age at birth and prevalence of asthma in a nationwide study of Latino children. METHODS: We included 3473 Latino children aged 8-21 years (1696 subjects with physician-diagnosed asthma and 1777 healthy controls) from five US centres and Puerto Rico recruited from July 2008 through November 2011. We used multiple logistic regression models to examine the effect of maternal age at birth on asthma in offspring overall and in analyses stratified by ethnic subgroup (Mexican American, Puerto Rican and other Latino). Secondary analyses evaluated the effects of siblings, acculturation and income on this relationship. RESULTS: Maternal age < 20 years was significantly associated with decreased odds of asthma in offspring, independent of other risk factors (OR = 0.73, 95% CI: 0.57-0.93). In subgroup analyses, the protective effect of younger maternal age was observed only in Mexican Americans (OR = 0.53, 95% CI: 0.36, 0.79). In Puerto Ricans, older maternal age was associated with decreased odds of asthma (OR = 0.65, 95% CI: 0.44-0.97). In further stratified models, the protective effect of younger maternal age in Mexican Americans was seen only in children without older siblings (OR = 0.44, 95% CI: 0.23-0.81). CONCLUSION AND CLINICAL RELEVANCE: In contrast to European descent populations, younger maternal age was associated with decreased odds of asthma in offspring in Mexican American women. Asthma is common in urban minority populations but the factors underlying the varying prevalence among different Latino ethnicities in the United States is not well understood. Maternal age represents one factor that may help to explain this variability.
Asunto(s)
Asma/epidemiología , Asma/etiología , Hispánicos o Latinos , Edad Materna , Adolescente , Estudios de Casos y Controles , Niño , Femenino , Hispánicos o Latinos/estadística & datos numéricos , Humanos , Masculino , Vigilancia de la Población , Factores de Riesgo , Estados Unidos/epidemiología , Adulto JovenRESUMEN
A hydrostatic pressure sensor based on morphology dependent resonances in a polymeric tube is presented. By internal pressurization, normal tensions will increase the device's size and shrink its wall thickness, inducing a shift in the resonant wavelengths of the resonator. Numerical simulations indicate that there are two modal regimes of sensitivity and a maximum achievable sensitivity, related to the device's geometry, constitutive material and analysed mode order. A sensitivity as high as 0.36 ± 0.01 nm/bar has been experimentally found for a 1.8mm diameter PMMA tube with wall thickness of 80µm.
RESUMEN
Hantavirus infection is an endemic zoonosis in Chile, with an average lethality of around 36%. The highest lethality (60%) was recorded in 1997. Prevention strategies have been applied since then. Early diagnosis and technologies, such as the use of ECMO (Extracorporeal Membrane Oxygenation) and Hantavirus immune plasma, have contributed to increasing the survival of people due to this disease at the national level. In the newly created Region of Ñuble in Chile, the incidence and lethality of Hantavirus cases are unknown; therefore, the objective of this research is to describe the epidemiological profile of Hantavirus cases in the Region of Ñuble, Chile from 2002 to 2018. This knowledge contributes to substantiating and justifies the need to invest in technology and reinforce interventions related to the early diagnosis and prevention of this disease in the region. Cases reported in the Ñuble region during the period 2002-2018, extracted from the Epidemiological Survey of Environmental Research of Hantavirus cases of the Ministry of Health of Chile, were analyzed retrospectively. The epidemiological profile of the Ñuble region is very similar to the national one in terms of characterizing the individual suffering from the disease. The most affected population is young men, residents in rural areas, and mainly from a low socioeconomic segment. The regional profile of Hantavirus cases makes it possible to identify three communes with the highest number of cases: El Carmen, Coihueco, and San Carlos. A political-administrative response is expected to focus on and optimize strategies and resources to reduce the incidence and lethality of this pathology in the Ñuble region.
Asunto(s)
Infecciones por Hantavirus , Orthohantavirus , Masculino , Animales , Humanos , Chile , Estudios Retrospectivos , ZoonosisRESUMEN
The photocatalytic efficiency of TiO(2)-SiMgO(x) plates to oxidize H(2)S was first evaluated in a flat laboratory reactor with 50 mL min(-1) synthetic air containing 100 ppm H(2)S in the presence of humidity. The use of the photocatalyst-adsorbent hybrid material enhanced the photocatalytic activity in terms of pollutant conversion, selectivity, and catalyst lifetime compared to previous H(2)S tests with pure TiO(2) because total H(2)S elimination was maintained for more than 30 operating hours with SO(2) appearing in the outlet as reaction product only after 18 h. Subsequently, the hybrid material was successfully tested in a photoreactor prototype to treat real polluted air in a wastewater treatment plant. For this purpose, a new tubular photocatalytic reactor that may use solar radiation in combination with artificial radiation was designed; the lamp was turned on when solar UV-A irradiance was below 20 W m(-2), which was observed to be the minimum value to ensure 100% conversion. The efficient distribution of the opaque photocatalyst inside the tubular reactor was achieved by using especially designed star-shaped structures. These structures were employed for the arrangement of groups of eight TiO(2)-SiMgO(x) plates in easy-to-handle channelled units obtaining an adequate flow regime without shading. The prototype continuously removed during one month and under real conditions the H(2)S contained in a 1 L min(-1) air current with a variable inlet concentration in the range of tens of ppmv without release of SO(2).
Asunto(s)
Contaminación del Aire/prevención & control , Sulfuro de Hidrógeno/efectos de la radiación , Fotólisis , Compuestos Orgánicos Volátiles/efectos de la radiación , Adsorción , Sulfuro de Hidrógeno/química , Silicatos de Magnesio/química , Titanio/química , Rayos Ultravioleta , Compuestos Orgánicos Volátiles/química , Purificación del AguaRESUMEN
In recent years, Aedes albopictus has become the most important invasive mosquito species worldwide. In 2018, Ae. albopictus was found in a suburban area of Merida, one of the cities with the highest number of arbovirus cases in Mexico in the last 10 years. As Ae. albopictus continues its range expansion, there is a need to monitor its susceptibility to existing insecticide classes, since countries like Mexico currently do not consider Ae. albopictus in its insecticide management programs. In order to determine its susceptibility to the insecticides usually applied by the vector control program in Mexico, the Centers for Disease Control and Prevention bottle bioassays were performed on individuals from established population of Ae. albopictus from Merida, Yucatan, Mexico. Results suggested that the population recently found in the suburban area of Merida is susceptible to permethrin, deltamethrin, chlorpyrifos, malathion, bendiocarb, and propoxur. Further studies of insecticide resistance using biochemical and molecular tools together with more knowledge of the biology and ecology of this species are necessary to generate specific and efficient control strategies in Mexico.
Asunto(s)
Aedes , Insecticidas , Animales , Humanos , Resistencia a los Insecticidas , Insecticidas/farmacología , México , Mosquitos VectoresRESUMEN
BACKGROUND: Leukotrienes play an important role in allergic and inflammatory diseases, but reports on the involvement of arachidonate 5-lipoxygenase-activating protein (ALOX5AP) and leukotriene A(4) hydrolase (LTA4H) in asthma have been inconclusive. OBJECTIVE: To determine whether polymorphisms in ALOX5AP and LTA4H genes are risk factors for asthma in two different Latino groups: Mexicans and Puerto Ricans. METHODS: The LTA4H gene was sequenced in individuals from both groups to identify novel polymorphisms. Single-nucleotide polymorphisms (SNPs) in the ALOX5AP and LTA4H genes were analysed for associations with asthma and asthma-related phenotypes in 687 parent-child trios of Mexican and Puerto Rican origin. RESULTS: In LTA4H, five previously unknown polymorphisms were identified. Two SNPs within LTA4H (rs17525488 and rs2540493) were protective for asthma in Latinos (P=0.007 and 0.05, respectively). Among the Mexican patients, LTA4H polymorphisms were associated with baseline lung function and IgE levels. For ALOX5AP, the minor allele at SNP rs10507391 was associated with protection from asthma (odds ratio=0.78, P=0.02) and baseline lung function (P=0.018) in Puerto Ricans. A gene-gene interaction was identified between LTA4H (rs17525488) and ALOX5AP (rs10507391), (P=0.003, in the combined sample). CONCLUSION: Our results support the role of LTA4H and ALOX5AP variants as risk factors for asthma in Latino populations.
Asunto(s)
Asma/genética , Proteínas Portadoras/genética , Epóxido Hidrolasas/genética , Predisposición Genética a la Enfermedad , Hispánicos o Latinos/genética , Proteínas de la Membrana/genética , Proteínas Activadoras de la 5-Lipooxigenasa , Adolescente , Alelos , Asma/etnología , Asma/fisiopatología , Proteínas Portadoras/metabolismo , Niño , Epóxido Hidrolasas/metabolismo , Femenino , Frecuencia de los Genes , Estudios de Asociación Genética , Humanos , Masculino , Proteínas de la Membrana/metabolismo , Americanos Mexicanos , Polimorfismo de Nucleótido Simple , Factores de Riesgo , Adulto JovenRESUMEN
Lake Laguna Santa Elena, a freshwater body, located in mid-south of Chile, is an environmental asset used as a water resource by the agricultural and touristic sector and is the habitat for a wide variety of endemic avifauna. The objective of this study was to assign a monetary value to this lacustrine body, using the methodology of AMUVAM (Multicriteria Analysis of Environmental Active Values). The information was collected through the application of surveys to a panel of experts involved in the use of the environmental resource. The result obtained with the AMUVAM method corresponds to the US $ 17,780,686, a figure that represents an estimated value of the TEV (Total Economic Value) of the environmental asset. This study, a pioneer in the Region, will support decision making, allowing an adequate management of this critical water resource.
Asunto(s)
Ecosistema , Lagos , Agricultura , Chile , Conservación de los Recursos NaturalesRESUMEN
Fluorescent carbon based-nanoparticles are one of the emerging nanomaterials. Their preparation is relatively simple, rapid and inexpensive, and they are less toxic compared with metal and semiconductor nanoparticles. Here, we report a simple and reliable method to prepare water-soluble fluorescent carbon nanoparticles (FC-NPs) from nanoparticles made from a protein, bovine serum albumin. The obtained mean size of our carbon nanoparticles is between 3.8 and 3.4â¯nm, and they exhibit its maximum fluorescence emission at 424 and 408â¯nm respectively (with a reasonable QY of 16.5%) due to the presence of functional groups (NH, NH2, COOH and OH) that contain O and N; the presence of these functional groups was confirmed by FTIR and XPS analysis. The photoluminescent decay lifetime was modeled by a two exponential fit which indicates a contribution from both core and surface states. Also, the preliminary results showed that FC-NPs had a good interaction with HeLa and normal oral epithelial cells; nanoparticles were permeable at the cell membrane and went to the cytosol, and even to the nucleus, in less than 30â¯min, the fluorescence images of our preliminary results did not show any apparent toxic damage in any of the cell lines.
Asunto(s)
Carbono/química , Colorantes Fluorescentes/química , Nanopartículas/química , Albúmina Sérica Bovina/química , Aminas/química , Animales , Ácidos Carboxílicos/química , Bovinos , Permeabilidad de la Membrana Celular , Células Epiteliales , Células HeLa , Humanos , Interacciones Hidrofóbicas e Hidrofílicas , Imagen Óptica , Espectrometría de Fluorescencia , Propiedades de SuperficieRESUMEN
The asthmatic response to the common cold is highly variable, and early characteristics that predict worsening of asthma control following a cold have not been identified. In this prospective multicentric cohort study of 413 adult subjects with asthma, the mini-Asthma Control Questionnaire (mini-ACQ) was used to quantify changes in asthma control and the Wisconsin Upper Respiratory Symptom Survey-21 (WURSS-21) to measure cold severity. Univariate and multivariable models were used to examine demographic, physiological, serological and cold-related characteristics for their relationship to changes in asthma control following a cold. Clinically significant worsening of asthma control was observed following a cold (mean+/-SD increase in mini-ACQ score of 0.69+/-0.93). Univariate analysis demonstrated that season, centre location, cold duration and cold severity measurements were all associated with a change in asthma control. Multivariable analysis of the covariates available within the first 2 days of cold onset revealed that the day 2 and cumulative sum of day 1 and 2 WURSS-21 scores were significant predictors of the subsequent changes in asthma control. In asthmatic subjects, cold severity within the first 2 days can be used to predict subsequent changes in asthma control. This information may help clinicians prevent deterioration in asthma control following a cold.
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Asma/diagnóstico , Asma/fisiopatología , Resfriado Común/complicaciones , Corticoesteroides/uso terapéutico , Adulto , Asma/etiología , Estudios de Cohortes , Femenino , Humanos , Masculino , Persona de Mediana Edad , Análisis Multivariante , Estudios Prospectivos , Calidad de Vida , Riesgo , Encuestas y Cuestionarios , Resultado del TratamientoRESUMEN
BACKGROUND: Eisenmenger syndrome (ES) is a heart cyanotic condition characterised by elevated pulmonary vascular resistance and an intra-cardiac right-to-left shunting of blood through a systemic-to-pulmonary circulation connection. Affected children usually exhibit severe hypoxia, clubbing of fingers/toes, haemoptysis, anaemia, and organ damage. CASE REPORT: During autumn 2015, the patient and her parents arrived at the paediatric dentistry clinic. The patient presented with the main complaint of generalised inflamed gingival tissues, severely protruded upper incisors, and evident abnormal mouth breathing. TREATMENT: This was performed under local analgesia, rubber-dam isolation, and antimicrobial prophylaxis with amoxicillin (50 mg/kg). The patient's parents agreed to the treatment plan through a signed informed consent. This treatment consisted of the placement of pit and fissure sealants on the four permanent first molars (which included enamel preparation with fissurotomy burs), in-depth gingiva/dental frequent cleanings, local fluoride varnish applications, and an exhaustive programme of at-home oral hygiene (brushing, flossing, and chlorhexidine mouth rinses), including adequate nutrition. Gingivoplasty surgery to remove residual enlarged tissues was indicated for the near future. FOLLOW-UP: The child did not return to the clinic. When contacted, the parents reported that their daughter's systemic condition worsened significantly. She was confined to a bed at home under palliative care, with a life-span expectation of only a few months. CONCLUSION: Comprehensive dental care of children with ES requires careful consideration of their medical condition, and dental care delivery should be coordinated with the paediatric cardiologist. General analgesia should be considered only in strictly selected cases, due to the high peri-operative mortality reported.
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Atención Dental para Niños/métodos , Complejo de Eisenmenger , Gingivitis/terapia , Anestesia Local , Niño , Caries Dental/complicaciones , Caries Dental/terapia , Complejo de Eisenmenger/complicaciones , Femenino , Gingivitis/complicaciones , Humanos , Respiración por la Boca/complicaciones , SobremordidaRESUMEN
In this work, gold nanospheres functionalized with low weight organic molecules (4-aminothiphenol and cysteamine) were synthesized in a one-step method for their in vitro cytotoxic evaluation on HeLa cells. To enhance the biocompatibility of the cysteamine-capped GNPs, BSA was used due to its broad PH stability and high binding affinity to gold nanoparticles. Besides, the widely reported silica coated gold nanorods were tested here to contrast their toxic response against our nanoparticles coated with organic molecules. Our results shown, the viability measured at 1.9×10-5M did not show significant differences against negative controls for all the samples; however, the metabolic activity of HeLa cells dropped when they were exposed to silica gold nanorods in the range of concentrations from 2.9×10-7M to 3.0×10-4M, while in the cases of gold nanospheres, we found that only at concentrations below 1.9×10-5M metabolic activity was normal. Our preliminary results did not indicate any perceivable harmful toxicity to cell membrane, cytoskeleton or nucleus due to our nanospheres at 1.9×10-5M. Additional test should be conducted in order to ensure a safe use of them for biological applications, and to determine the extent of possible damage.
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Oro/toxicidad , Nanopartículas del Metal/toxicidad , Compuestos de Anilina/química , Supervivencia Celular/efectos de los fármacos , Cisteamina/química , Citoesqueleto/efectos de los fármacos , Oro/química , Células HeLa , Humanos , Nanopartículas del Metal/química , Nanotubos/química , Nanotubos/toxicidad , Albúmina Sérica Bovina/química , Dióxido de Silicio/química , Compuestos de Sulfhidrilo/químicaRESUMEN
Substitution of amino acids within the section of Tn21 resolvase that corresponds to a helix-turn-helix structure, with the equivalent residues from Tn3 resolvase, yields proteins that retain the ability to mediate recombination between res sites from Tn21. These proteins had no recombinational activity on res sites from Tn3, even when the complete recognition helix had been exchanged. In this study, the binding of these mutants of Tn21 resolvase to DNA fragments containing res from either Tn21 or Tn3 was analysed by DNase I footprinting and by gel retardation. With DNA containing res from Tn21, the mutants bound to all three of the binding sites for resolvase (I, II, and III) but with a lower affinity than wild-type Tn21 resolvase. No complexes were detected between Tn3 resolvase and Tn21 DNA. With DNA containing res from Tn3, both the mutants and wild-type Tn21 resolvase bound to sites II and III, forming similar complexes to those with Tn3 resolvase: some of the mutants had higher affinities for these two sites on Tn3 DNA than on Tn21 DNA. In contrast, at site I in res from Tn3 (the location of the recombinational cross-over), the derivatives of Tn21 resolvase formed aberrant complexes whose structures differed radically from that with Tn3 resolvase. Alterations in the amino acid sequence of resolvase, within the helix-turn-helix region, therefore modulate the affinity of the protein for its target sequence in the DNA, but the specificity of resolvase for recombination at its cognate res sites is determined by the resultant organization of the DNA-protein complex.
Asunto(s)
Proteínas de Unión al ADN/metabolismo , Nucleotidiltransferasas/metabolismo , Resolvasas de Transposones , Secuencia de Bases , ADN/metabolismo , Elementos Transponibles de ADN/genética , Desoxirribonucleasa I , Escherichia coli/enzimología , Mutagénesis , Nucleotidiltransferasas/genética , Proteínas Recombinantes de Fusión/metabolismo , Recombinación Genética/fisiología , TransposasasRESUMEN
Plasmid R6K contains two functional origins or transfer (oriT), in contrast to previously characterized conjugative plasmids. The oriTs are formed by 98 bp palindromic sequences invertedly orientated with respect to each other and located in the immediate vicinity of the alpha and beta origins of replication. The gene for R6K oriT-nickase, taxC, was identified by transposon mutagenesis and sequenced, revealing that TaxC belongs to the VirD2 nickase family. The protein was overproduced and purified. It catalysed a cleaving-joining reaction on single-stranded DNA containing its target sequence. Identification of the nic sites suggested that the R6K oriTs belong to the RP4/VirD2 oriT family. Cleavage was highly specific and did not occur with oligonucleotides cleaved by related nickases like TraI of RP4 or VirD2 of the Ti plasmid. nic cleavage of in vivo preassembled relaxation complexes was induced by incubation of plasmid cleared lysates with ethidium bromide. Nicked molecules obtained in this way were treated with snake venom phosphodiesterase to produce double strand cleavages at the nic sites. 35% of the molecules were cleaved simultaneously at both nic sites, both in the case of R6K and of R6Kdrd1, a derepressed mutant whose frequency of transfer is 1000-fold higher. This figure represents the minimum percentage of individual R6K molecules containing two pre-assembled relaxation complexes.
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Conjugación Genética/genética , Proteínas de Unión al ADN/genética , Plásmidos/genética , Secuencia de Aminoácidos , Secuencia de Bases , Clonación Molecular , ADN Bacteriano/metabolismo , ADN de Cadena Simple/metabolismo , Proteínas de Unión al ADN/biosíntesis , Proteínas de Unión al ADN/aislamiento & purificación , Proteínas de Unión al ADN/metabolismo , Escherichia coli/enzimología , Escherichia coli/genética , Genes Bacterianos/genética , Datos de Secuencia Molecular , Peso Molecular , Oligodesoxirribonucleótidos/metabolismo , Hidrolasas Diéster Fosfóricas , Plásmidos/metabolismo , Origen de Réplica , Análisis de Secuencia de ADNRESUMEN
The resolvases from the transposons Tn3 and Tn21 are homologous proteins but they possess distinct specificities for the DNA sequence at their respective res sites. The DNA binding domain of resolvase contains an amino acid sequence that can be aligned with the helix-turn-helix motif of other DNA binding proteins. Mutations in the gene for Tn21 resolvase were made by replacing the section of DNA that codes for the helix-turn-helix with synthetic oligonucleotides. Each mutation substituted one amino acid in Tn21 resolvase with either the corresponding residue from Tn3 resolvase or a residue that lacks hydrogen bonding functions. The ability of these proteins to mediate recombination between res sites from either Tn21 or Tn3 was measured in vivo and in vitro. With one exception, where a glutamate residue had been replaced by leucine, the activity of these mutants was similar to that of wild-type Tn21 resolvase. A further mutation was made in which the complete recognition helix of Tn21 resolvase was replaced with that from Tn3 resolvase. This protein retained activity in recombining Tn21 res sites, though at a reduced level relative to wild-type; the reduction can be assigned entirely to weakened binding to this DNA. Neither this mutant nor any other derivative of Tn21 resolvase had any detectable activity for recombination between res sites from Tn3. The exchange of this section of amino acid sequence between the two resolvases is therefore insufficient to alter the DNA sequence specificity for recombination.
Asunto(s)
Nucleotidiltransferasas/metabolismo , Recombinación Genética/fisiología , Resolvasas de Transposones , Secuencia de Aminoácidos , Secuencia de Bases , Elementos Transponibles de ADN , Proteínas de Unión al ADN/genética , Escherichia coli/enzimología , Datos de Secuencia Molecular , Mutagénesis , Nucleotidiltransferasas/genética , Proteínas Recombinantes de Fusión/metabolismo , Relación Estructura-Actividad , TransposasasRESUMEN
Protein TrwC is the relaxase-helicase responsible for the initiation and termination reactions of DNA processing during plasmid R388 conjugation. Site-directed mutagenesis was used to change to phenylalanine each of a set of four conserved tyrosyl residues in the sequence of the N-terminal relaxation domain of the protein. Simultaneous mutation of both Y18 and Y26 was required to abolish in vitro cleavage and strand-transfer reactions catalyzed by protein TrwC on oligonucleotides containing the nic site. Thus, both Y18 and Y26 could be involved independently in the formation of oligonucleotide-protein covalent complexes that constitute presumed intermediates of these reactions. This hypothesis was confirmed by the observation of Y18 and Y26-specific peptide-oligonucleotide adducts after protease digestion of TrwC and mutant derivatives. Finally mutation Y18F, but not mutation Y26F, abolished nic-cleavage of a supercoiled DNA containing the R388 origin of transfer (oriT). These data allowed the construction of a model for conjugative DNA processing in which Y18 specifically catalyzes the initial cleavage reaction, while Y26 is used for the second strand-transfer reaction, which terminates conjugation. The model suggests a control mechanism that can be effective at each conjugative replication cycle.
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Conjugación Genética/genética , ADN Nucleotidiltransferasas/química , ADN Nucleotidiltransferasas/metabolismo , Escherichia coli/enzimología , Integrasas , Plásmidos/genética , Tirosina/metabolismo , Secuencia de Aminoácidos , Sustitución de Aminoácidos/genética , Sitios de Unión , Catálisis , Aductos de ADN/química , Aductos de ADN/genética , Aductos de ADN/metabolismo , ADN Nucleotidiltransferasas/genética , ADN Superhelicoidal/genética , Endopeptidasas/metabolismo , Escherichia coli/genética , Modelos Genéticos , Datos de Secuencia Molecular , Mutagénesis Sitio-Dirigida/genética , Oligodesoxirribonucleótidos/genética , Fragmentos de Péptidos/química , Fragmentos de Péptidos/genética , Fragmentos de Péptidos/metabolismo , Recombinasas , Secuencias Reguladoras de Ácidos Nucleicos/genética , Alineación de Secuencia , Factores de Tiempo , Tirosina/genéticaRESUMEN
Abstract Lake Laguna Santa Elena, a freshwater body, located in mid-south of Chile, is an environmental asset used as a water resource by the agricultural and touristic sector and is the habitat for a wide variety of endemic avifauna. The objective of this study was to assign a monetary value to this lacustrine body, using the methodology of AMUVAM (Multicriteria Analysis of Environmental Active Values). The information was collected through the application of surveys to a panel of experts involved in the use of the environmental resource. The result obtained with the AMUVAM method corresponds to the US $ 17,780,686, a figure that represents an estimated value of the TEV (Total Economic Value) of the environmental asset. This study, a pioneer in the Region, will support decision making, allowing an adequate management of this critical water resource.
Resumo A Laguna Santa Elena, um corpo de água doce localizado no centro-sul do Chile, é um ativo ambiental usado como recurso hídrico pelos setores agrícola e turístico e é o habitat de uma ampla variedade de avifauna endêmica. O objetivo deste estudo foi atribuir um valor monetário a esse corpo lacustre, utilizando a metodologia da AMUVAM (Análise Multicritério de Valores Ambientais Ativos). As informações foram coletadas por meio da aplicação de pesquisas a um painel de especialistas envolvidos no uso do recurso ambiental. O resultado obtido com o método AMUVAM corresponde a US$ 17.780.686, que representam um valor estimado do valor econômico total (VT) do ativo ambiental. Este estudo, pioneiro na região, apoiará a tomada de decisões, permitindo um gerenciamento adequado desse recurso hídrico crítico.