RESUMO
Complete genome sequencing was performed for Anabaena variabilis ATCC 29413 from the collection of the Chair of Genetics, Department of Biology, Moscow State University, Russia. In addition to known plasmids A, B, and C, a new circular low-copy plasmid was detected and named D. It was also sequenced completely and found to have 27051 bp. The plasmid contained the parA and parB genes of the partition system, two genes that encode replication proteins, a gene for site-specific recombinase, atype-I restriction-modification system, and several genes with unknown functions. Analysis by PCR revealed the presence of plasmid D in two epiphytic strains from Vietnam, i.e., Anabaena sp. 182 and Anabaena sp. 281, as well as in Anabaena sp. V5 and A. azollae (Newton's isolate).
Assuntos
Anabaena variabilis/genética , Plasmídeos/genética , Anabaena variabilis/isolamento & purificação , DNA Nucleotidiltransferases , Genes Bacterianos , Análise de Sequência de DNA , VietnãRESUMO
A gene encoding superoxide dismutase was revealed in the genome of the thermoacidophilic crenarchaeon Acidilobus saccharovorans. A recombinant expression vector was constructed and transformed into E. coli cells. The novel recombinant superoxide dismutase was purified and characterized. The enzyme was shown to be an iron-dependent superoxide dismutase able to bind various bivalent metals in the active site. According to differential scanning calorimetric data, the denaturation temperature of the enzyme is 107.3°C. The maximal activity of the Fe(II) reconstituted enzyme defined by xanthine oxidase assay is 1700 U/mg protein. Study of the thermal stability of the superoxide dismutase samples with various metal contents by tryptophan fluorescence indicated that the thermal stability and activity of the enzyme directly depend on the nature of the reconstituted metal and the degree of saturation of binding sites.
Assuntos
Crenarchaeota/enzimologia , Superóxido Dismutase/genética , Superóxido Dismutase/metabolismo , Sequência de Aminoácidos , Ativação Enzimática , Estabilidade Enzimática , Escherichia coli/genética , Fontes Termais/microbiologia , Concentração de Íons de Hidrogênio , Multimerização Proteica , Estrutura Quaternária de Proteína , Superóxido Dismutase/química , Superóxido Dismutase/isolamento & purificação , Superóxidos/metabolismo , TemperaturaRESUMO
Prolidases are peptidases that are specific for dipeptides with proline as the second residue. The structure of recombinant prolidase from the hyperthermophilic archaeon Thermococcus sibiricus (Tsprol) was determined at 2.6â Å resolution. The homodimer of Tsprol is characterized by a complete lack of interactions between the N- and C-terminal domains of the two subunits and hence can be considered to be the most open structure when compared with previously structurally studied prolidases. This structure exists owing to intermolecular coordination bonds between cadmium ions derived from the crystallization solution and histidine residues of a His tag and aspartate and glutamate residues, which link the dimers to each other. This linking leads to the formation of a crystal with a loose packing of protein molecules and low resistance to mechanical influence and temperature increase.
Assuntos
Proteínas Arqueais/química , Dipeptidases/química , Thermococcus/enzimologia , Domínio Catalítico , Cristalografia por Raios X , Modelos Moleculares , Domínios e Motivos de Interação entre Proteínas , Estrutura Quaternária de Proteína , Estrutura Secundária de Proteína , Subunidades Proteicas/química , Proteínas Recombinantes/químicaRESUMO
As a result of sequencing the genome of the termophilic alkali-tolerant lipolytic bacterium Thermosyntropha lipolytica, the gene encoding a lipase secreted into the medium was identified. The recombinant enzyme was expressed in Escherichia coli. It was isolated, purified, and functionally characterized. The lipase exhibited hydrolytic activity toward para-nitrophenyl esters of various chain lengths, as well as triglycerides, including vegetable oils. The optimal reaction conditions were achieved at temperatures from 70 to 80 degrees C and pH 8.0. Enzyme saved more than 80% of its activity in the presence of 10% methanol. This new thermostable lipase may be a promising biocatalyst for organic synthesis; it may find application in the food and detergent industry and biodiesel production.
Assuntos
Proteínas de Bactérias/genética , Genoma Bacteriano , Bactérias Gram-Positivas Formadoras de Endosporo/enzimologia , Lipase/genética , Óleos de Plantas/metabolismo , Álcalis , Sequência de Aminoácidos , Proteínas de Bactérias/isolamento & purificação , Proteínas de Bactérias/metabolismo , Clonagem Molecular , Escherichia coli , Bactérias Gram-Positivas Formadoras de Endosporo/genética , Temperatura Alta , Concentração de Íons de Hidrogênio , Lipase/isolamento & purificação , Lipase/metabolismo , Lipólise , Dados de Sequência Molecular , Nitrofenóis , Proteínas Recombinantes/genética , Proteínas Recombinantes/isolamento & purificação , Proteínas Recombinantes/metabolismo , Triglicerídeos/metabolismoAssuntos
Aminopeptidases/metabolismo , Desulfurococcaceae/enzimologia , Aminopeptidases/química , Aminopeptidases/genética , Aminopeptidases/isolamento & purificação , Clonagem Molecular , Desulfurococcaceae/genética , Cinética , Metais/farmacologia , Multimerização Proteica , Estrutura Quaternária de ProteínaAssuntos
Archaea/classificação , Cianobactérias/classificação , Fontes Termais/microbiologia , Archaea/genética , Archaea/isolamento & purificação , Cianobactérias/genética , Cianobactérias/isolamento & purificação , DNA Ribossômico/genética , RNA Ribossômico 16S/genética , Federação Russa , Análise de Sequência de DNA , Enxofre/metabolismoRESUMO
Alcohol dehydrogenases belong to the oxidoreductase family and play an important role in a broad range of physiological processes. They catalyze the cofactor-dependent reversible oxidation of alcohols to the corresponding aldehydes or ketones. The NADP-dependent short-chain alcohol dehydrogenase TsAdh319 from the thermophilic archaeon Thermococcus sibiricus was overexpressed, purified and crystallized. Crystals were obtained using the hanging-drop vapour-diffusion method using 25%(w/v) polyethylene glycol 3350 pH 7.5 as precipitant. The crystals diffracted to 1.68 A resolution and belonged to space group I222, with unit-cell parameters a = 55.63, b = 83.25, c = 120.75 A.
