RESUMO
High-gravity (HG) brewing has broader application to succeed on beer differentiation and production optimization. However, such process imposes a handicap to yeasts, which must be able to deal with stressful conditions in fermentation. In this work, we assessed different physiological traits of 24 Saccharomyces cerevisiae strains isolated from Brazilian bioethanol distilleries for the selection of novel starters for HG brewing. Five yeast strains were selected with ability to overcome different stressors under HG beer fermentation, showing high fermentability rates, resilience to ethanol stress, low production of foam and hydrogen sulfide, as well as similar flocculation rates to brewer's yeasts. After five fermentation recycles, most strains sustained a viability rate higher than 90% and were able to efficiently accumulate trehalose and glycogen, besides presenting no detectable petite mutants at the final stage. In the sensory analysis, the beers obtained from the five selected strains showed greater aromatic complexity, with predominance of 'spicy', 'dried' and 'fresh fruits' descriptors. In conclusion, this study sheds light on the potential of yeast strains from Brazilian bioethanol process to produce distinctive specialty beers, aside from proposing an effective selection methodology based on relevant physiological attributes for HG brewing process.
Assuntos
Hipergravidade , Saccharomyces cerevisiae , Cerveja , Brasil , FermentaçãoRESUMO
An early step of target validation in antimicrobial drug discovery is to prove that a gene coding for a putative target is essential for pathogen's viability. However, little attention has been paid to demonstrate the causal links between gene essentiality and a particular protein function that will be the focus of a drug discovery effort. This should be considered an important step in target validation since a growing number of proteins are found to exhibit multiple and unrelated tasks. Here, we show that the Mycobacterium tuberculosis (Mtb) folB gene is essential and that this essentiality depends on the dihydroneopterin aldolase/epimerase activities of its protein product, the FolB protein from the folate biosynthesis pathway. The wild-type (WT) MtFolB and point mutants K99A and Y54F were cloned, expressed, purified and monitored for the aldolase, epimerase and oxygenase activities using HPLC. In contrast to the WT MtFolB, both mutants have neither aldolase nor epimerase activities in the conditions assayed. We then performed gene knockout experiments and showed that folB gene is essential for Mtb survival under the conditions tested. Moreover, only the WT folB sequence could be used as a rescue copy in gene complementation studies. When the sequences of mutants K99A or Y54F were used for complementation, no viable colonies were obtained, indicating that aldolase and/or epimerase activities are crucial for Mtb survival. These results provide a solid basis for further work aiming to develop new anti-TB agents acting as inhibitors of the aldolase/epimerase activities of MtFolB.
Assuntos
Aldeído Liases/antagonistas & inibidores , Antituberculosos/farmacologia , Proteínas de Bactérias/antagonistas & inibidores , Descoberta de Drogas/métodos , Mycobacterium tuberculosis/efeitos dos fármacos , Aldeído Liases/genética , Aldeído Liases/metabolismo , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Biocatálise , Cromatografia Líquida de Alta Pressão , Genes Essenciais/genética , Teste de Complementação Genética/métodos , Humanos , Viabilidade Microbiana/efeitos dos fármacos , Viabilidade Microbiana/genética , Terapia de Alvo Molecular/métodos , Mutação de Sentido Incorreto , Mycobacterium tuberculosis/enzimologia , Mycobacterium tuberculosis/genética , Reprodutibilidade dos Testes , Especificidade por Substrato , Espectrometria de Massas em Tandem , Tuberculose/tratamento farmacológico , Tuberculose/microbiologiaRESUMO
Feedbacks between land carbon pools and climate provide one of the largest sources of uncertainty in our predictions of global climate. Estimates of the sensitivity of the terrestrial carbon budget to climate anomalies in the tropics and the identification of the mechanisms responsible for feedback effects remain uncertain. The Amazon basin stores a vast amount of carbon, and has experienced increasingly higher temperatures and more frequent floods and droughts over the past two decades. Here we report seasonal and annual carbon balances across the Amazon basin, based on carbon dioxide and carbon monoxide measurements for the anomalously dry and wet years 2010 and 2011, respectively. We find that the Amazon basin lost 0.48 ± 0.18 petagrams of carbon per year (Pg C yr(-1)) during the dry year but was carbon neutral (0.06 ± 0.1 Pg C yr(-1)) during the wet year. Taking into account carbon losses from fire by using carbon monoxide measurements, we derived the basin net biome exchange (that is, the carbon flux between the non-burned forest and the atmosphere) revealing that during the dry year, vegetation was carbon neutral. During the wet year, vegetation was a net carbon sink of 0.25 ± 0.14 Pg C yr(-1), which is roughly consistent with the mean long-term intact-forest biomass sink of 0.39 ± 0.10 Pg C yr(-1) previously estimated from forest censuses. Observations from Amazonian forest plots suggest the suppression of photosynthesis during drought as the primary cause for the 2010 sink neutralization. Overall, our results suggest that moisture has an important role in determining the Amazonian carbon balance. If the recent trend of increasing precipitation extremes persists, the Amazon may become an increasing carbon source as a result of both emissions from fires and the suppression of net biome exchange by drought.
Assuntos
Atmosfera/química , Ciclo do Carbono , Secas/estatística & dados numéricos , Biomassa , Biota , Brasil , Dióxido de Carbono/análise , Monóxido de Carbono/análise , Incêndios/estatística & dados numéricos , Água Doce/análise , Fotossíntese , Chuva , Estações do Ano , Árvores/metabolismo , Clima TropicalRESUMO
Blood pressure (BP) and physical activity (PA) levels are inversely associated. Since genetic factors account for the observed variation in each of these traits, it is possible that part of their association may be related to common genetic and/or environmental influences. Thus, this study was designed to estimate the genetic and environmental correlations of BP and PA phenotypes in nuclear families from Muzambinho, Brazil. Families including 236 offspring (6 to 24 years) and their 82 fathers and 122 mothers (24 to 65 years) were evaluated. BP was measured, and total PA (TPA) was assessed by an interview (commuting, occupational, leisure time, and school time PA). Quantitative genetic modeling was used to estimate maximal heritability (h²), and genetic and environmental correlations. Heritability was significant for all phenotypes (systolic BP: h² = 0.37 ± 0.10, P < 0.05; diastolic BP: h² = 0.39 ± 0.09, P < 0.05; TPA: h² = 0.24 ± 0.09, P < 0.05). Significant genetic (r g) and environmental (r e) correlations were detected between systolic and diastolic BP (r g = 0.67 ± 0.12 and r e = 0.48 ± 0.08, P < 0.05). Genetic correlations between BP and TPA were not significant, while a tendency to an environmental cross-trait correlation was found between diastolic BP and TPA (r e = -0.18 ± 0.09, P = 0.057). In conclusion, BP and PA are under genetic influences. Systolic and diastolic BP share common genes and environmental influences. Diastolic BP and TPA are probably under similar environmental influences.
Assuntos
Adolescente , Adulto , Idoso , Criança , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Adulto Jovem , Pressão Sanguínea/genética , Interação Gene-Ambiente , Predisposição Genética para Doença/genética , Hipertensão/genética , Atividade Motora/genética , Brasil , Característica Quantitativa HerdávelRESUMO
Blood pressure (BP) and physical activity (PA) levels are inversely associated. Since genetic factors account for the observed variation in each of these traits, it is possible that part of their association may be related to common genetic and/or environmental influences. Thus, this study was designed to estimate the genetic and environmental correlations of BP and PA phenotypes in nuclear families from Muzambinho, Brazil. Families including 236 offspring (6 to 24 years) and their 82 fathers and 122 mothers (24 to 65 years) were evaluated. BP was measured, and total PA (TPA) was assessed by an interview (commuting, occupational, leisure time, and school time PA). Quantitative genetic modeling was used to estimate maximal heritability (h²), and genetic and environmental correlations. Heritability was significant for all phenotypes (systolic BP: h² = 0.37 ± 0.10, P < 0.05; diastolic BP: h² = 0.39 ± 0.09, P < 0.05; TPA: h² = 0.24 ± 0.09, P < 0.05). Significant genetic (r g) and environmental (r e) correlations were detected between systolic and diastolic BP (r g = 0.67 ± 0.12 and r e = 0.48 ± 0.08, P < 0.05). Genetic correlations between BP and TPA were not significant, while a tendency to an environmental cross-trait correlation was found between diastolic BP and TPA (r e = -0.18 ± 0.09, P = 0.057). In conclusion, BP and PA are under genetic influences. Systolic and diastolic BP share common genes and environmental influences. Diastolic BP and TPA are probably under similar environmental influences.
Assuntos
Pressão Sanguínea/genética , Interação Gene-Ambiente , Predisposição Genética para Doença/genética , Hipertensão/genética , Atividade Motora/genética , Adolescente , Adulto , Idoso , Brasil , Criança , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Característica Quantitativa Herdável , Adulto JovemRESUMO
Septins are evolutionarily conserved proteins that contain a GTPase domain and are capable of forming filaments at the cell periphery. Septins are involved in many essential cellular processes, such as cytokinesis and cell polarization, and are used as markers of morphogenesis in several fungi. Dimorphism in fungi enables cells to switch between morphologies (yeast or filament forms), due to changes in the temperature of the environment. We analysed the localization of septin proteins in yeast and filamentous cells of the dimorphic fungus Paracoccidioides brasiliensis, a common cause of granulomatous mycosis. In order to determine septin localization, we first cloned Cdc12p, a septin homolog from P. brasiliensis, and expressed it in Escherichia coli. Following PbCdc12p purification, specific serum against PbCdc12p were raised for use in immunofluorescence assays. We observed the hourglass and ring forms of septin filaments during cell division in yeast. Septin filaments were also simultaneously localized in the necks of multiple budding cells. A distinctive pattern of punctuate and/or diffuse localization was also seen in the periphery of multinucleate yeast cells and at the tips and septa of filamentous cells. A more diffuse and punctuate pattern of localization observed in P. brasiliensis cells seems to be unique to filamentous and dimorphic fungi and may be related to their specialization in cell wall deposition, morphogenesis and cell cycle control.
Assuntos
Proteínas Fúngicas/análise , Paracoccidioides/metabolismo , Septinas/análise , Divisão Celular , Escherichia coli/genética , Imunofluorescência , Proteínas Fúngicas/química , Proteínas Fúngicas/genética , Hifas/metabolismo , Paracoccidioides/ultraestrutura , Filogenia , Septinas/química , Septinas/genéticaRESUMO
The objective was to evaluate the effect of slaughter weight on the processing efficiency of Pacu (Piaractus mesopotamicus).The fish were cultivated in a soil vivarium and fed commercial feed. After ten months of cultivation, 96 fish were deadened, slaughtered, weighed and dissected to determine the yield of clean body, fillet and percentage of entrails. The weight classes analyzed were: CI - 135 to 285g; CII - 310 to 385g; CIII - 400 to 585g; CIV - 600 to 1.285g. For the variance analysis the data was grouped in four weight classes, and the regression equations were estimated regarding mean weight in each class. No effect of the weight class (P>0,05) was observed on the yield of clean body (RTL). However, there was a linear effect (P<0,05) of the weight classes in slaughter over the fillet yield (RF) and percentage of entrails. Pacus slaughtered in more elevated weight classes had greater filled yield. Thus, the fish destined to the fillet process should preferably be slaughtered at higher weights. The fish cultivated with the objective of being commercialized as a carcass or whole with entrails removed may be slaughtered at lower weights.
Assuntos
Animais , Indústria Pesqueira , Água Doce , Peixes/crescimento & desenvolvimento , Manipulação de AlimentosRESUMO
The enzymes of the shikimate pathway represent potential molecular targets for the development of non-toxic antimicrobial agents and anti-parasite drugs. One of the most promising of these enzymes is shikimate kinase (EC 2.7.1.71), which is responsible for the fifth step in the shikimate pathway. This enzyme phosphorylates shikimic acid to yield shikimate-3-phosphate, using ATP as a substrate. In this work, the conformational dynamics of the shikimate kinase from Mycobacterium tuberculosis was investigated in its apostate in solution. For this study, the enzyme was subjected to a gradient of temperatures from 15°C to 45°C in the presence or absence of deuterium oxide, and the amide H/D exchange was monitored using ESI-mass spectrometry. We observed: i) the phosphate binding domain in the apo-enzyme is fairly rigid and largely protected from solvent access, even at relatively high temperatures; ii) the shikimate binding domain is highly flexible, as indicated by the tendency of the apo-enzyme to exhibit large conformational changes to permit LID closure after the shikimate binding; iii) the nucleotide binding domain is initially conformationally rigid, which seems to favour the initial orientation of ADP/ATP, but becomes highly flexible at temperatures above 30°C, which may permit domain rotation; iv) part of the LID domain, including the phosphate binding site, is partially rigid, while another part is highly flexible and accessible to the solvent.
Assuntos
Antituberculosos/farmacologia , Mycobacterium tuberculosis/enzimologia , Fosfotransferases (Aceptor do Grupo Álcool)/química , Sequência de Aminoácidos , Antituberculosos/química , Antituberculosos/uso terapêutico , Medição da Troca de Deutério , Óxido de Deutério/química , Inibidores Enzimáticos/química , Inibidores Enzimáticos/farmacologia , Humanos , Cinética , Dados de Sequência Molecular , Fosfotransferases (Aceptor do Grupo Álcool)/antagonistas & inibidores , Fosfotransferases (Aceptor do Grupo Álcool)/metabolismo , Ligação Proteica , Estrutura Secundária de Proteína , Estrutura Terciária de Proteína , Espectrometria de Massas por Ionização por Electrospray , Temperatura , Tuberculose/tratamento farmacológicoRESUMO
The causative agent of tuberculosis (TB), Mycobacterium tuberculosis, infects one-third of the world population. TB remains the leading cause of mortality due to a single bacterial pathogen. The worldwide increase in incidence of M. tuberculosis has been attributed to the high proliferation rates of multi and extensively drug-resistant strains, and to co-infection with the human immunodeficiency virus. There is thus a continuous requirement for studies on mycobacterial metabolism to identify promising targets for the development of new agents to combat TB. Singular characteristics of this pathogen, such as functional and structural features of enzymes involved in fundamental metabolic pathways, can be evaluated to identify possible targets for drug development. Enzymes involved in the pyrimidine salvage pathway might be attractive targets for rational drug design against TB, since this pathway is vital for all bacterial cells, and is composed of enzymes considerably different from those present in humans. Moreover, the enzymes of the pyrimidine salvage pathway might have an important role in the mycobacterial latent state, since M. tuberculosis has to recycle bases and/or nucleosides to survive in the hostile environment imposed by the host. The present review describes the enzymes of M. tuberculosis pyrimidine salvage pathway as attractive targets for the development of new antimycobacterial agents. Enzyme functional and structural data have been included to provide a broader knowledge on which to base the search for compounds with selective biological activity.
Assuntos
Mycobacterium tuberculosis/enzimologia , Pirimidinas/metabolismo , Citidina Desaminase/metabolismo , Mycobacterium tuberculosis/metabolismo , Núcleosídeo-Difosfato Quinase/metabolismo , Núcleosídeo-Fosfato Quinase/metabolismo , Nucleotídeo Desaminases/metabolismo , Pentosiltransferases/metabolismo , Pirimidina Fosforilases/metabolismo , Pirofosfatases/metabolismo , Timidilato Sintase/metabolismoRESUMO
Millions of deaths worldwide are caused by the aetiological agent of tuberculosis, Mycobacterium tuberculosis. The increasing prevalence of this disease, the emergence of drug-resistant strains, and the devastating effect of human immunodeficiency virus coinfection have led to an urgent need for the development of new and more efficient antimycobacterial drugs. The modern approach to the development of new chemical compounds against complex diseases, especially the neglected endemic ones, such as tuberculosis, is based on the use of defined molecular targets. Among the advantages, this approach allows (i) the search and identification of lead compounds with defined molecular mechanisms against a specific target (e.g. enzymes from defined pathways), (ii) the analysis of a great number of compounds with a favorable cost/benefit ratio, and (iii) the development of compounds with selective toxicity. The present review describes the enzymes of the purine salvage pathway in M. tuberculosis as attractive targets for the development of new antimycobacterial agents. Enzyme kinetics and structural data have been included to provide a thorough knowledge on which to base the search for compounds with biological activity. We have focused on the mycobacterial homologues of this pathway as potential targets for the development of new antitubercular agents.
Assuntos
Mycobacterium tuberculosis/enzimologia , Purinas/metabolismo , 5'-Nucleotidase/metabolismo , Adenosina Desaminase/metabolismo , Adenosina Quinase/metabolismo , Adenilossuccinato Liase/metabolismo , Adenilossuccinato Sintase/metabolismo , IMP Desidrogenase/metabolismo , Mycobacterium tuberculosis/metabolismo , N-Glicosil Hidrolases/metabolismo , Núcleosídeo-Fosfato Quinase/metabolismo , Pentosiltransferases/metabolismo , Purina-Núcleosídeo Fosforilase/metabolismoRESUMO
The objective was to evaluate the effect of slaughter weight on the processing efficiency of Pacu (Piaractus mesopotamicus).The fish were cultivated in a soil vivarium and fed commercial feed. After ten months of cultivation, 96 fish were deadened, slaughtered, weighed and dissected to determine the yield of clean body, fillet and percentage of entrails. The weight classes analyzed were: CI - 135 to 285g; CII - 310 to 385g; CIII - 400 to 585g; CIV - 600 to 1.285g. For the variance analysis the data was grouped in four weight classes, and the regression equations were estimated regarding mean weight in each class. No effect of the weight class (P>0,05) was observed on the yield of clean body (RTL). However, there was a linear effect (P<0,05) of the weight classes in slaughter over the fillet yield (RF) and percentage of entrails. Pacus slaughtered in more elevated weight classes had greater filled yield. Thus, the fish destined to the fillet process should preferably be slaughtered at higher weights. The fish cultivated with the objective of being commercialized as a carcass or whole with entrails removed may be slaughtered at lower weights.(AU)
Assuntos
Animais , Peixes/crescimento & desenvolvimento , Água Doce , Indústria Pesqueira , Manipulação de AlimentosRESUMO
BACKGROUND AND OBJECTIVE: Purine nucleoside phosphorylase (PNP) is an enzyme that catalyzes the reversible phosphorolysis of purine nucleosides, playing a key role in the purine salvage pathway. Activated T cells seem to rely heavily on PNP to remain functionally active and are particularly sensitive to PNP deficiency. The role of PNP in periodontal tissues has not been characterized thus far. The aim of this study therefore was to assess the activity and expression of PNP in the gingival tissues of periodontitis patients. MATERIAL AND METHODS: Ten patients consecutively admitted for treatment had their periodontal clinical variables recorded and their gingival crevicular fluid collected. After periodontal treatment the patients were seen once a month for plaque and bleeding control, and had their periodontal variables recorded and gingival crevicular fluid collected at 90 and 180 d. Purine nucleoside phosphorylase-specific activity was assessed using a spectrophotometer through the addition of the PNP substrate analog 2-amino-6mercapto-7-methyl purine riboside to the gingival crevicular fluid. In parallel, PNP expression was assessed by immunohistochemistry and real-time PCR in gingival biopsies and cell culture. RESULTS: Purine nucleoside phosphorylase activity was higher in the gingival crevicular fluid of periodontally diseased sites, which was positively correlated with improvements of the clinical variables. Treatment of periodontal disease induced a striking decrease of PNP activity in periodontally diseased sites. Expression of PNP was more pronounced in mononuclear cells and endothelial cells of the gingiva, and the mRNA levels were 5.7-fold higher in inflamed tissues compared with control samples. CONCLUSION: Purine nucleoside phosphorylase activity and expression are upregulated in periodontally diseased sites and can be detected in the gingival crevicular fluid.
Assuntos
Periodontite Agressiva/enzimologia , Periodontite Crônica/enzimologia , Líquido do Sulco Gengival/enzimologia , Purina-Núcleosídeo Fosforilase/genética , Purina-Núcleosídeo Fosforilase/metabolismo , Adulto , Idoso , Periodontite Agressiva/terapia , Linfócitos T CD4-Positivos/enzimologia , Periodontite Crônica/terapia , Regulação Enzimológica da Expressão Gênica , Gengiva/enzimologia , Humanos , Memória Imunológica , Pessoa de Meia-Idade , Distribuição Normal , Estatísticas não Paramétricas , Regulação para CimaRESUMO
AIMS: To evaluate the dominance and persistence of strains of Saccharomyces cerevisiae during the process of sugar cane fermentation for the production of cachaça and to analyse the microbial compounds produced in each fermentative process. METHODS AND RESULTS: Three S. cerevisiae strains were evaluated during seven consecutive 24-h fermentation batches using recycled inocula. The UFLA CA 116 strain had the largest population of viable organisms, and the maximum population was achieved in the fourth batch after 96 h of fermentation. The UFLA CA 1162 and UFLA CA 1183 strains grew more slowly, and the maximum population was reached in the seventh batch. Molecular characterization of isolated yeast cells using PFGE (pulse field gel electrophoresis) revealed that more than 86% of the isolates corresponded to the initially inoculated yeast strain. The concentration of aldehydes, esters, methanol, alcohol and volatile acids in the final-aged beverages were within the legal limits. CONCLUSIONS: Cachaça produced by select yeast strains exhibits analytical differences. UFLA CA 1162 and UFLA CA 116 S. cerevisiae isolates can be considered the ideal strains for the artisanal production of cachaça in Brazil. SIGNIFICANCE AND IMPACT OF THE STUDY: The use of select yeast strains can improve the quality and productivity of cachaça production. Our findings are important for the appropriate monitoring of yeast during sugar cane fermentation. In addition, we demonstrate that UFLA CA 116 and UFLA CA 1162, the ideal yeast strains for cachaça production, are maintained at a high population density. The persistence of these yeast strains in the fermentation of sugar cane juice promotes environmental conditions that prevent or decrease bacterial contamination. Thus, the use of select yeast strains for the production of cachaça is a viable economic alternative to standardize the production of this beverage.
Assuntos
Bebidas Alcoólicas/microbiologia , Fermentação , Microbiologia de Alimentos , Saccharomyces cerevisiae/crescimento & desenvolvimento , Brasil , Cariotipagem , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/isolamento & purificação , Saccharum/microbiologiaRESUMO
This work describes for the first time a model of Purine Nucleoside Phosphorylase from Listeria monocytogenes (LmPNP). We modeled the complexes of LmPNP with ligands in order to determine the structural basis for specificity. Comparative analysis of the model of LmPNP allowed identification of structural features responsible for ligand affinities.
Assuntos
Biologia Computacional , Listeria monocytogenes/enzimologia , Purina-Núcleosídeo Fosforilase/química , Sequência de Aminoácidos , Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , Apoenzimas/antagonistas & inibidores , Apoenzimas/química , Apoenzimas/metabolismo , Sítios de Ligação , Desenho de Fármacos , Humanos , Ligantes , Listeria monocytogenes/efeitos dos fármacos , Listeriose/tratamento farmacológico , Modelos Moleculares , Estrutura Terciária de Proteína , Purina-Núcleosídeo Fosforilase/antagonistas & inibidores , Purina-Núcleosídeo Fosforilase/metabolismo , Especificidade por SubstratoRESUMO
The rate at which knowledge about genomic sequences and their protein products is produced is increasing much faster than the rate of 3-dimensional protein structure determination by experimental methods, such as X-ray diffraction and nuclear magnetic resonance. One of the major challenges in structural bioinformatics is the conversion of genomic sequences into useful information, such as characterization of protein structure and function. Using molecular dynamics (MD) simulations, we predicted the 3-dimensional structure of an artificially designed three- alpha -helix bundle, called A3, from a fully extended initial conformation, based on its amino acid sequence. The MD protocol enabled us to obtain the secondary, in 1.0 ns, as well as the supersecondary and tertiary structures, in 4.0-10.0 ns, of A3, much faster than previously described for a similar protein system. The structure obtained at the end of the 10.0-ns MD simulation was topologically a three-alpha-helix bundle.
Assuntos
Simulação por Computador , Modelos Moleculares , Proteínas/química , Sequência de Aminoácidos , Dados de Sequência Molecular , Estrutura Secundária de Proteína , SolventesRESUMO
Tuberculosis (TB) and Malaria are neglected diseases, which continue to be major causes of morbidity and mortality worldwide, killing together around 5 million people each year. Mycolic acids, the hallmark of mycobacteria, are high-molecular-weight alpha-alkyl, beta-hydroxy fatty acids. Biochemical and genetic experimental data have shown that the product of the M. tuberculosis inhA structural gene (InhA) is the primary target of isoniazid mode of action, the most prescribed anti-tubercular agent. InhA was identified as an NADH-dependent enoyl-ACP(CoA) reductase specific for long-chain enoyl thioesters and is a member of the Type II fatty acid biosynthesis system, which elongates acyl fatty acid precursors of mycolic acids. M. tuberculosis and P. falciparum enoyl reductases are targets for the development of anti-tubercular and antimalarial agents. Here we present a brief description of the mechanism of action of, and resistance to, isoniazid. In addition, data on inhibition of mycobacterial and plasmodial enoyl reductases by triclosan are presented. We also describe recent efforts to develop inhibitors of M. tuberculosis and P. falciparum enoyl reductase enzyme activity.
Assuntos
Antimaláricos/administração & dosagem , Antituberculosos/administração & dosagem , Sistemas de Liberação de Medicamentos/métodos , Desenho de Fármacos , Enoil-(Proteína de Transporte de Acila) Redutase (NADH)/metabolismo , Animais , Antimaláricos/síntese química , Antituberculosos/síntese química , Sistemas de Liberação de Medicamentos/tendências , Enoil-(Proteína de Transporte de Acila) Redutase (NADH)/antagonistas & inibidores , Enoil-(Proteína de Transporte de Acila) Redutase (NADH)/química , Inibidores Enzimáticos/administração & dosagem , Inibidores Enzimáticos/síntese química , HumanosRESUMO
Purine nucleoside phosphorylase (PNP) catalyzes the reversible phosphorolysis of nucleosides and deoxynucleosides, generating ribose 1-phosphate and the purine base, which is an important step of purine catabolism pathway. The lack of such an activity in humans, owing to a genetic disorder, causes T-cell impairment, and thus drugs that inhibit human PNP activity have the potential of being utilized as modulators of the immunological system to treat leukemia, autoimmune diseases, and rejection in organ transplantation. Besides, the purine salvage pathway is the only possible way for apicomplexan parasites to obtain the building blocks for RNA and DNA synthesis, which makes PNP from these parasites an attractive target for drug development against diseases such as malaria. Hence, a number of research groups have made efforts to elucidate the mechanism of action of PNP based on structural and kinetic studies. It is conceivable that the mechanism may be different for PNPs from diverse sources, and influenced by the oligomeric state of the enzyme in solution. Furthermore, distinct transition state structures can make possible the rational design of specific inhibitors for human and apicomplexan enzymes. Here, we review the current status of these research efforts to elucidate the mechanism of PNP-catalyzed chemical reaction, focusing on the mammalian and Plamodium falciparum enzymes, targets for drug development against, respectively, T-Cell- and Apicomplexan parasites-mediated diseases.
Assuntos
Apicomplexa/enzimologia , Sistemas de Liberação de Medicamentos/métodos , Infecções por Protozoários/enzimologia , Purina-Núcleosídeo Fosforilase/metabolismo , Linfócitos T/enzimologia , Animais , Apicomplexa/patogenicidade , Humanos , Infecções por Protozoários/tratamento farmacológico , Infecções por Protozoários/parasitologia , Purina-Núcleosídeo Fosforilase/antagonistas & inibidores , Linfócitos T/parasitologiaRESUMO
The aetiological agent of tuberculosis (TB), Mycobacterium tuberculosis, is responsible for millions of deaths annually. The increasing prevalence of the disease, the emergence of multidrug-resistant strains, and the devastating effect of human immunodeficiency virus co-infection have led to an urgent need for the development of new and more efficient antimycobacterial drugs. Since the shikimate pathway is present and essential in algae, higher plants, bacteria, and fungi, but absent from mammals, the gene products of the common pathway might represent attractive targets for the development of new antimycobacterial agents. In this review we describe studies on shikimate pathway enzymes, including enzyme kinetics and structural data. We have focused on mycobacterial shikimate pathway enzymes as potential targets for the development of new anti-TB agents.
Assuntos
Sistemas de Liberação de Medicamentos/métodos , Desenho de Fármacos , Mycobacterium tuberculosis/enzimologia , Ácido Chiquímico/metabolismo , Transdução de Sinais/efeitos dos fármacos , Animais , Inibidores Enzimáticos/administração & dosagem , Inibidores Enzimáticos/síntese química , Humanos , Mycobacterium tuberculosis/efeitos dos fármacos , Ácido Chiquímico/antagonistas & inibidores , Ácido Chiquímico/síntese química , Transdução de Sinais/fisiologiaRESUMO
The rate at which knowledge about genomic sequences and their protein products is produced is increasing much faster than the rate of 3-dimensional protein structure determination by experimental methods, such as X-ray diffraction and nuclear magnetic resonance. One of the major challenges in structural bioinformatics is the conversion of genomic sequences into useful information, such as characterization of protein structure and function. Using molecular dynamics (MD) simulations, we predicted the 3-dimensional structure of an artificially designed three-alpha-helix bundle, called A3, from a fully extended initial conformation, based on its amino acid sequence. The MD protocol enabled us to obtain the secondary, in 1.0 ns, as well as the supersecondary and tertiary structures, in 4.0-10.0 ns, of A3, much faster than previously described for a similar protein system. The structure obtained at the end of the 10.0-ns MD simulation was topologically a three-alpha-helix bundle.
Assuntos
Simulação por Computador , Modelos Moleculares , Proteínas/química , Sequência de Aminoácidos , Dados de Sequência Molecular , Estrutura Secundária de Proteína , SolventesRESUMO
The DNA puff BhC4-1 gene of the sciarid Bradysia hygida is induced in salivary glands prior to the pupal molt as a secondary response to the increase in ecdysone titers. Previous studies demonstrated that the BhC4-1 promoter is activated in transgenic Drosophila melanogaster salivary glands as a late response to the ecdysone peak that triggers metamorphosis, revealing that this aspect of BhC4-1 transcriptional regulation is conserved in the Drosophila background. To identify regulators of BhC4-1 expression, we utilized a candidate gene approach and tested the roles of the ecdysone-induced genes BR-C, E74, and E75. Our results reveal that the BR-C Z3 isoform is essential for BhC4-1-lacZ induction in prepupal salivary glands and constitute the first demonstration of the participation of early genes products on DNA puff genes regulation.