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Your search keyword '"Vara, Y"' showing total 4 results
Start Over Author "Vara, Y" Topic genes, bacterial
4 results on '"Vara, Y"'

1. Silencing of the gene coding for the epsilon subunit of DNA polymerase III slows down the growth rate of Escherichia coli populations.

Author

Stefan A, Reggiani L, Cianchetta S, Radeghieri A, Gonzalez Vara y Rodriguez A, and Hochkoeppler A

Subjects
Base Sequence, Blotting, Northern, Cloning, Molecular, Exodeoxyribonuclease V, Molecular Sequence Data, Nucleic Acid Conformation, RNA, Antisense chemistry, DNA Polymerase III, Escherichia coli growth & development, Escherichia coli Proteins, Exodeoxyribonucleases genetics, Gene Silencing, Genes, Bacterial
Abstract

Chromosome replication in Escherichia coli is accomplished by the multimeric enzyme DNA polymerase III; the relevance, in vivo, of the epsilon subunit (encoded by dnaQ) for processivity and fidelity of DNA polymerase III has been evaluated. To this aim, dnaQ has been conditionally silenced by means of in vivo expression of different antisense RNAs. Unexpectedly, the presence of the Shine-Dalgarno sequence is essential for the effectiveness of antisense constructs. Silencing of dnaQ induces a severe decrease in growth rate not paralleled by high mutation frequencies, suggesting that the epsilon subunit primarily affects the processivity of DNA polymerase III.

Published
2003
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2. Nucleotide sequence, expression and transcriptional analysis of the Bifidobacterium longum MB 219 lacZ gene.

Author

Rossi M, Altomare L, Gonzàlez Vara y Rodriguez A, Brigidi P, and Matteuzzi D

Subjects
Base Sequence, Bifidobacterium enzymology, Cloning, Molecular, DNA Primers genetics, DNA, Bacterial genetics, Escherichia coli genetics, Gene Expression, Species Specificity, Transcription, Genetic, beta-Galactosidase genetics, Bifidobacterium genetics, Genes, Bacterial, Lac Operon
Abstract

The gene encoding beta-galactosidase was isolated by functional complementation of Escherichia coli from Bifidobacterium longum MB219, which exhibited the highest activity among ten Bifidobacterium strains tested of the species B. longum, B. breve, B. adolescentis, B. indicum, B. animalis and B. cuniculi. The nucleotide sequence of the 5.0-kb fragment conferring the positive beta-galactosidase phenotype to E. coli revealed the presence of a lacZ-type gene encoding a 1023-amino-acid protein that was preceded by a ribosome binding site. A sequence showing 72% identity with the proline tRNA of Bacillus subtilis and a gene probably encoding the DNA-3-methyladenine glycosydase I were located downstream from the lacZ gene, after a gap of 30-50 unsequenced base pairs. By primer-extension analysis, the transcription start site of the lacZ gene was mapped 65 nt upstream from the start codon, and it enabled identification of the -10 region of the putative promoter. The nucleotide sequence of lacZ and its deduced amino acid sequence were compared with those of beta-galactosidase genes and enzymes from other microorganisms. High similarity was demonstrated between the B. longum beta-galactosidase and its counterparts in Lactobacillus delbruckii subsp. bulgaricus, Streptococcus salivarius subsp. thermophilus, E. coli, Clostridium acetobutylicum, Leuconostoc lactis, Klebsiella pneumoniae and Kluyveromyces marxianus var. lactis, all belonging to the LacZ family. The B. longum MB219 lacZ gene was cloned in Bifidobacterium and its expression was observed in strains with otherwise low levels of endogenous activity. The expression increased by factors of 1.5-50 and enabled those strains that do not grow on lactose to use this sugar as sole carbon source.

Published
2000
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3. Directed evolution of β-galactosidase from Escherichia coli by mutator strains defective in the 3′→5′ exonuclease activity of DNA polymerase III

Author

Alejandro Hochkoeppler, Alessandra Stefan, Annalisa Radeghieri, and Antonio Gonzalez Vara y Rodriguez

Subjects
Exonuclease, Exodeoxyribonuclease V, DNA polymerase, DNA polymerase II, β-Galactosidase, Biophysics, medicine.disease_cause, Biochemistry, Structural Biology, Escherichia coli, Genetics, medicine, Molecular Biology, Conjugative plasmid, Polymerase, DNA Polymerase III, DNA Primers, Base Sequence, biology, beta-Glucosidase, Mutagenesis, Cell Biology, beta-Galactosidase, Directed evolution, Molecular biology, Exodeoxyribonucleases, Lac Operon, Genes, Bacterial, biology.protein, 3'-5' Exonuclease, Mutator strain, Directed Molecular Evolution
Abstract

Directed evolution of Escherichia coli beta-galactosidase into variants featuring beta-glucosidase activity was challenged. To this end, mutagenesis of lacZ was performed by replication in E. coli CC954, a mutator strain containing a DNA polymerase III defective in 3'--5' exonuclease activity. beta-Galactosidase variants can be isolated upon mutagenesis of lacZ hosted into the self-transmissible episome F'128. Optimal evolution of lacZ can be achieved by propagation of E. coli CC954/F'128 cultures for 15 generations; further growth of mutator cultures for 37 or 55 generations imposes a high mutational load on lacZ and hinders the selection of efficiently evolved clones.

Published
2001
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4. Nucleotide sequence, expression and transcriptional analysis of the Bifidobacterium longum MB 219 lacZ gene

Author

Luigi Altomare, Patrizia Brigidi, Diego Matteuzzi, Gonzàlez Vara y Rodriguez A, and Maddalena Rossi

Subjects
DNA, Bacterial, Bifidobacterium longum, Transcription, Genetic, Gene Expression, medicine.disease_cause, Biochemistry, Microbiology, Species Specificity, Gene expression, Escherichia coli, Genetics, medicine, Cloning, Molecular, Molecular Biology, Peptide sequence, Gene, LacZ, Expression, Transcriptional analysis, Nucleotide sequence, DNA Primers, Base Sequence, biology, Nucleic acid sequence, food and beverages, General Medicine, beta-Galactosidase, biology.organism_classification, Ribosomal binding site, Complementation, Lac Operon, Genes, Bacterial, Bifidobacterium
Abstract

The gene encoding beta-galactosidase was isolated by functional complementation of Escherichia coli from Bifidobacterium longum MB219, which exhibited the highest activity among ten Bifidobacterium strains tested of the species B. longum, B. breve, B. adolescentis, B. indicum, B. animalis and B. cuniculi. The nucleotide sequence of the 5.0-kb fragment conferring the positive beta-galactosidase phenotype to E. coli revealed the presence of a lacZ-type gene encoding a 1023-amino-acid protein that was preceded by a ribosome binding site. A sequence showing 72% identity with the proline tRNA of Bacillus subtilis and a gene probably encoding the DNA-3-methyladenine glycosydase I were located downstream from the lacZ gene, after a gap of 30-50 unsequenced base pairs. By primer-extension analysis, the transcription start site of the lacZ gene was mapped 65 nt upstream from the start codon, and it enabled identification of the -10 region of the putative promoter. The nucleotide sequence of lacZ and its deduced amino acid sequence were compared with those of beta-galactosidase genes and enzymes from other microorganisms. High similarity was demonstrated between the B. longum beta-galactosidase and its counterparts in Lactobacillus delbruckii subsp. bulgaricus, Streptococcus salivarius subsp. thermophilus, E. coli, Clostridium acetobutylicum, Leuconostoc lactis, Klebsiella pneumoniae and Kluyveromyces marxianus var. lactis, all belonging to the LacZ family. The B. longum MB219 lacZ gene was cloned in Bifidobacterium and its expression was observed in strains with otherwise low levels of endogenous activity. The expression increased by factors of 1.5-50 and enabled those strains that do not grow on lactose to use this sugar as sole carbon source.

Published
2000

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