RESUMEN
The NADPH-dependent homodimeric flavoenzyme thioredoxin reductase (TrxR) provides reducing equivalents to thioredoxin, a key regulator of various cellular redox processes. Crystal structures of photo-inactivated thioredoxin reductase (TrxR) from the Gram-positive bacterium Lactococcus lactis have been determined. These structures reveal novel molecular features that provide further insight into the mechanisms behind the sensitivity of this enzyme toward visible light. We propose that a pocket on the si-face of the isoalloxazine ring accommodates oxygen that reacts with photo-excited FAD generating superoxide and a flavin radical that oxidize the isoalloxazine ring C7α methyl group and a nearby tyrosine residue. This tyrosine and key residues surrounding the oxygen pocket are conserved in enzymes from related bacteria, including pathogens such as Staphylococcus aureus. Photo-sensitivity may thus be a widespread feature among bacterial TrxR with the described characteristics, which affords applications in clinical photo-therapy of drug-resistant bacteria.
Asunto(s)
Lactococcus lactis/enzimología , Lactococcus lactis/efectos de la radiación , Luz , Estrés Oxidativo , Procesos Fotoquímicos , Reductasa de Tiorredoxina-Disulfuro/química , Reductasa de Tiorredoxina-Disulfuro/metabolismo , Flavina-Adenina Dinucleótido/metabolismo , Flavinas/química , Flavinas/metabolismo , Redes y Vías Metabólicas , Modelos Moleculares , Conformación Molecular , Oxidación-Reducción , Relación Estructura-ActividadRESUMEN
Lactococcus lactis is a culture widely used in salt-containing dairy products. Salt hinders bacterial growth, but exposure to environmental stress may protect cells against subsequent stress, including salt. The objective of this study was to evaluate the salt tolerance of L. lactis R-604 after exposure to various stresses. The culture was subjected to 10% (vol/vol) ethanol for 30 min, mild heat at 52°C for 30 min, 15 mM hydrogen peroxide for 30 min, or UV light (254 nm) for 5 min and compared with a control. Starting with 5 log cfu/mL for all treatments, growth was determined in M17 broth with 5 NaCl concentrations (0, 1, 3, 5, and 7% wt/vol). Plating was conducted daily for 5 d. Salt tolerance was enhanced with mild heat exposure before growth in M17 broth with 5% (wt/vol) NaCl on d 3, 4, and 5, and with exposure to hydrogen peroxide and ethanol stresses before growth in M17 broth with 5% (wt/vol) NaCl on d 4 and 5. Exposure of this culture to mild heat, hydrogen peroxide, or ethanol before growth in M17 broth containing 5% (wt/vol) salt can enhance its survival, which could be beneficial when using it in salt-containing dairy products.
Asunto(s)
Lactococcus lactis/fisiología , Tolerancia a la Sal/fisiología , Estrés Fisiológico , Animales , Medios de Cultivo , Etanol/farmacología , Calor , Peróxido de Hidrógeno/farmacología , Lactococcus lactis/efectos de los fármacos , Lactococcus lactis/efectos de la radiación , Tolerancia a la Sal/efectos de los fármacos , Tolerancia a la Sal/efectos de la radiación , Estrés Fisiológico/efectos de los fármacos , Estrés Fisiológico/efectos de la radiación , Rayos UltravioletaRESUMEN
Metabolome analysis and physicochemical analyses were executed with cell extracts of a Lactococcus lactis subspecies cremoris strain treated by moderate pulsed electric field (PEF) to elucidate the mechanism of enhanced production of exopolysaccharide (EPS) by the treatment. Metabolome analysis by capillary electrophoresis time of flight mass spectrometry annotated 224 metabolites from the cytoplasmic extract of the strain, which, however, showed no significant changes in metabolites related to the EPS production. Electron microscopic observation and chemical analysis of undecaprenoids as carrier of EPS biosynthetic intermediates suggested that PEF treatment dissociated immature EPSs from the intermediates due to the focal electro-condensation of hydrogen ions at the cell surface. Thus, liberated undecaprenyl phosphates were recycled efficiently, which resulted in mass increase of EPS with smaller molecular weight. The study suggested the feasibility of moderate PEF treatment as a food processing technique and revealed the mechanism of enhanced production of EPS by the treatment.
Asunto(s)
Lactococcus lactis/química , Metaboloma/genética , Polisacáridos Bacterianos/biosíntesis , Animales , Vías Biosintéticas/genética , Citoplasma/química , Citoplasma/genética , Campos Electromagnéticos , Concentración de Iones de Hidrógeno , Hidrólisis , Lactococcus lactis/efectos de la radiación , Microscopía Electrónica , Peso Molecular , Fosfatos de Poliisoprenilo/química , Polisacáridos Bacterianos/químicaRESUMEN
OBJECTIVES: Lycopene biosynthetic genes from Deinococcus radiodurans were co-expressed in Lactococcus lactis to produce lycopene and improve its tolerance to stress. RESULTS: Lycopene-related genes from D. radiodurans, DR1395 (crtE), DR0862 (crtB), and DR0861 (crtI), were fused in line with S hine-Dalgarno (SD) sequences and co-expressed in L. lactis. The recombinant strain produced 0.36 mg lycopene g-1 dry cell wt after 48 h fermentation. The survival rate to UV irradiation of the recombinant strain was higher than that of the non-transformed strain. CONCLUSION: The L. lactis with co-expressed genes responsible for lycopene biosynthesis from D. radiodurans produced lycopene and exhibited increased resistance to UV stress, suggesting that the recombinant strain has important application potential in food industry.
Asunto(s)
Carotenoides/metabolismo , Deinococcus/genética , Lactococcus lactis/metabolismo , Fermentación , Lactococcus lactis/genética , Lactococcus lactis/efectos de la radiación , Licopeno , Rayos UltravioletaRESUMEN
Lactococcus lactis MG1363 was found to be unable to grow at temperatures above 37°C in a defined medium without riboflavin, and the cause was identified to be dissolved oxygen introduced during preparation of the medium. At 30°C, growth was unaffected by dissolved oxygen and oxygen was consumed quickly. Raising the temperature to 37°C resulted in severe growth inhibition and only slow removal of dissolved oxygen. Under these conditions, an abnormally low intracellular ratio of [ATP] to [ADP] (1.4) was found (normally around 5), which indicates that the cells are energy limited. By adding riboflavin to the medium, it was possible to improve growth and oxygen consumption at 37°C, and this also normalized the [ATP]-to-[ADP] ratio. A codon-optimized redox-sensitive green fluorescent protein (GFP) was introduced into L. lactis and revealed a more oxidized cytoplasm at 37°C than at 30°C. These results indicate that L. lactis suffers from heat-induced oxidative stress at increased temperatures. A decrease in intracellular flavin adenine dinucleotide (FAD), which is derived from riboflavin, was observed with increasing growth temperature, but the presence of riboflavin made the decrease smaller. The drop was accompanied by a decrease in NADH oxidase and pyruvate dehydrogenase activities, both of which depend on FAD as a cofactor. By overexpressing the riboflavin transporter, it was possible to improve FAD biosynthesis, which resulted in increased NADH oxidase and pyruvate dehydrogenase activities and improved fitness at high temperatures in the presence of oxygen.
Asunto(s)
Lactococcus lactis/fisiología , Lactococcus lactis/efectos de la radiación , Estrés Oxidativo , Riboflavina/metabolismo , Adenosina Difosfato/análisis , Adenosina Trifosfato/biosíntesis , Medios de Cultivo/química , Flavina-Adenina Dinucleótido/análisis , Genes Reporteros , Proteínas Fluorescentes Verdes/análisis , Proteínas Fluorescentes Verdes/genética , Cetona Oxidorreductasas/metabolismo , Lactococcus lactis/crecimiento & desarrollo , Lactococcus lactis/metabolismo , Complejos Multienzimáticos/metabolismo , NADH NADPH Oxidorreductasas/metabolismo , TemperaturaRESUMEN
Bacteria are normally haploid, maintaining one copy of their genome in one circular chromosome. We have examined the cell cycle of laboratory strains of Lactococcus lactis, and, to our surprise, we found that some of these strains were born with two complete nonreplicating chromosomes. We determined the cellular content of DNA by flow cytometry and by radioactive labeling of the DNA. These strains thus fulfill the criterion of being diploid. Several dairy strains were also found to be diploid while a nondairy strain and several other dairy strains were haploid in slow-growing culture. The diploid and haploid strains differed in their sensitivity toward UV light, in their cell size, and in their D period, the period between termination of DNA replication and cell division.
Asunto(s)
Cromosomas Bacterianos , Diploidia , Lactococcus lactis/genética , ADN Bacteriano/análisis , Citometría de Flujo/métodos , Lactococcus lactis/química , Lactococcus lactis/efectos de la radiación , Trazadores Radiactivos , Coloración y Etiquetado/métodos , Rayos UltravioletaRESUMEN
AIMS: The aim of this study was to isolate bacteriocin-producing lactic acid bacteria (LAB) from human intestine. METHODS AND RESULTS: A total of 111 LAB were isolated from human adult stool and screened for their bacteriocin production. Neutralized cell-free supernatants from Lactococcus lactis subsp. lactis MM19 and Pediococcus acidilactici MM33 showed antimicrobial activity. The antimicrobials in the supernatant from a culture of L. lactis inhibited Enterococcus faecium, various species of Lactobacillus and Staphylococcus aureus; while those in the supernatant from a culture of P. acidilactici inhibited Enterococcus spp., some lactobacilli and various serotypes of Listeria monocytogenes. The antimicrobial metabolites were heat-stable and were active over a pH range of 2-10. The antimicrobial activities of the supernatants of both bacteria were inhibited by many proteases but not by catalase. The plate overlay assay allowed an approximation of size between 3.5 and 6 kDa for both antimicrobial substances. CONCLUSIONS: As the antagonistic factor(s) produced by L. lactis MM19 and P. acidilactici MM33 were sensitive to proteolytic enzymes, it could be hypothesized that bacteriocins were involved in the inhibitory activities. Inhibition spectrum and biochemical analysis showed that these bacteria produced two distinct bacteriocins. SIGNIFICANCE AND IMPACT OF THE STUDY: We are the first to isolate bacteriocin-producing strains of Pediococcus and Lactococcus from human intestine. These strains might be useful for control of enteric pathogens.
Asunto(s)
Antibacterianos/farmacología , Bacteriocinas/farmacología , Lactococcus lactis/metabolismo , Pediococcus/metabolismo , Antibacterianos/metabolismo , Proteínas Bacterianas/análisis , Bacteriocinas/metabolismo , Medios de Cultivo , Electroforesis en Gel de Poliacrilamida/métodos , Enterococcus/efectos de los fármacos , Heces/microbiología , Rayos gamma , Calor , Humanos , Concentración de Iones de Hidrógeno , Intestinos/microbiología , Lactobacillus/efectos de los fármacos , Lactococcus lactis/aislamiento & purificación , Lactococcus lactis/efectos de la radiación , Listeria monocytogenes/efectos de los fármacos , Pediococcus/aislamiento & purificación , Pediococcus/efectos de la radiación , Staphylococcus aureus/efectos de los fármacos , Factores de TiempoRESUMEN
To increase the nisin synthesizing activity of the natural strain Lactococcus lactis subsp. lactis 119 isolated from sour milk, UV irradiation in different doses was used followed by isolation of productive clones. The highest mutation effect was observed with the dose of 76,000 erg/mm2, when 11% of the cells increased their productivity by 12.8% at the minimum survival rate. Two-step UV irradiation and adaptive selection on the nisin-contaning medium provided isolation of a strain with the activity 42.6% higher than that of the initial strain (3850 IU/ml). Natural and UV-induced variability of the strain by the nisin synthesis, growth rate, carbohydrate consumption and sensitivity to antibiotics of various groups were studied.
Asunto(s)
Lactococcus lactis/efectos de la radiación , Nisina/biosíntesis , Rayos Ultravioleta , Animales , Lactococcus lactis/metabolismo , Viabilidad Microbiana/efectos de la radiaciónRESUMEN
Fluorescence correlation spectroscopy (FCS) under two-photon excitation was applied successfully to characterize the penetration and diffusion capabilities of fluorescent probes (latex beads and fluorescein isothiocyanate-dextran) of different size and electrical charge in two models of monomicrobial biofilms with low (Lactococcus lactis biofilm) or high (Stenotrophonas maltophilia biofilm) contents of extracellular polymeric substance (EPS). FCS measurements performed on each biofilm can show deviation from Brownian diffusion, depending on the local structure of the biofilm and the fluorophore size. In this case, we fitted the data to an anomalous diffusion model and determined apparent diffusion coefficients, which can be 50 times smaller than the values in aqueous solutions. This result was interpreted as steric hindrance of the diffusion of the fluorescent particles within the biofilm that can lead to a total inhibition as observed particularly in the mushroom-like structure of the S. maltophilia biofilm. Alternatively, mechanisms for the absence of FCS signal behavior were related to attractive electrostatic interactions between cationic particles and negatively charged bacteria or to specific interactions between dextrans and EPS of the biofilm matrix.
Asunto(s)
Bacterias/crecimiento & desarrollo , Biopelículas/crecimiento & desarrollo , Fotones , Bacterias/efectos de la radiación , Bacterias/ultraestructura , Biopelículas/efectos de la radiación , Difusión , Colorantes Fluorescentes , Lactococcus lactis/crecimiento & desarrollo , Lactococcus lactis/efectos de la radiación , Lactococcus lactis/ultraestructura , Microscopía Electrónica de Rastreo , Modelos Biológicos , Espectrometría de FluorescenciaRESUMEN
The regulatory functions of the leader region preceding the Lactococcus lactis trp operon have been studied by mutagenesis analysis. This leader presents striking similarity to 'T-box' leaders found upstream of many Gram-positive aminoacyl-tRNA synthetase genes and some amino acid biosynthesis operons, which are controlled by antitermination through interaction of the leader transcript with cognate uncharged tRNA. A region of the L. lactis leader transcript also contains a series of (G/U) AG repeats which, in Bacillus, are involved in the binding of the trp RNA-binding protein (TRAP) which controls trp transcription. A screen was developed for the isolation of regulatory mutants affected in the leader region. All spontaneous mutants contained deletions; point mutations were only obtained after UV-induced mutagenesis. All mutations affected the putative transcription terminator upstream of the trp operon, demonstrating that trp is indeed controlled by transcription antitermination.
Asunto(s)
Lactococcus lactis/genética , Operón/genética , Transcripción Genética/genética , Triptófano/genética , Secuencia de Bases , Northern Blotting , Codón de Terminación/genética , Codón de Terminación/efectos de la radiación , Lactococcus lactis/crecimiento & desarrollo , Lactococcus lactis/efectos de la radiación , Datos de Secuencia Molecular , Mutación/genética , Mutación/efectos de la radiación , Operón/efectos de la radiación , ARN/análisis , Secuencias Reguladoras de Ácidos Nucleicos/genética , Secuencias Reguladoras de Ácidos Nucleicos/efectos de la radiación , Análisis de Secuencia de ARN , Transcripción Genética/efectos de la radiación , Triptófano/biosíntesis , Triptófano/efectos de la radiación , Rayos UltravioletaRESUMEN
The aim of the present study was to investigate the influence of 308 nm excimer-laser radiation on bacterial growth. Six different bacterial strains (Staphylococcus aureus, Escherichia coli, Streptococcus faecalis, Lactococcis lactis, Salmonella typhimurium, and Deinococcus radiodurans) were exposed in vitro to various doses and energy densities of laser radiation. To exclude bacterial killing by supraphysiological heating, the temperature change in the samples during irradiation was measured. Extended antimicrobial effects of XeCl excimer-laser radiation depending on the time of radiation, the energy density of the laser beam, and the irradiated bacterial strain were observed. Reduction of bacterial growth is independent of temperature and not linked to any ablative tissue removal. In almost all cases, a 99.9% reduction of bacteria was reached by total radiation times < 100 ms. The proven antimicrobial effects of 308 nm excimer-laser radiation may be of significant clinical importance in endodontics and periodontology in the future.
Asunto(s)
Bacterias/efectos de la radiación , Rayos Láser , Análisis de Varianza , Enterococcus faecalis/efectos de la radiación , Escherichia coli/efectos de la radiación , Cocos Grampositivos/efectos de la radiación , Lactococcus lactis/efectos de la radiación , Terapia por Láser , Gases Nobles , Fenómenos Físicos , Física , Salmonella typhimurium/efectos de la radiación , Staphylococcus aureus/efectos de la radiación , Temperatura , Factores de Tiempo , XenónRESUMEN
Studies of cellular responses to DNA-damaging agents, mostly in Escherichia coli, have revealed numerous genes and pathways involved in DNA repair. However, other species, particularly those which exist under different environmental conditions than does E. coli, may have rather different responses. Here, we identify and characterize genes involved in DNA repair in a gram-positive plant and dairy bacterium, Lactococcus lactis. Lactococcal strain MG1363 was mutagenized with transposition vector pG+host9::ISS1, and 18 mutants sensitive to mitomycin and UV were isolated at 37 degrees C. DNA sequence analyses allowed the identification of 11 loci and showed that insertions are within genes implicated in DNA metabolism (polA, hexB, and deoB), cell envelope formation (gerC and dltD), various metabolic pathways (arcD, bglA, gidA, hgrP, metB, and proA), and, for seven mutants, nonidentified open reading frames. Seven mutants were chosen for further characterization. They were shown to be UV sensitive at 30 degrees C (the optimal growth temperature of L. lactis); three (gidA, polA, and uvs-75) were affected in their capacity to mediate homologous recombination. Our results indicate that UV resistance of the lactococcal strain can be attributed in part to DNA repair but also suggest that other factors, such as cell envelope composition, may be important in mediating resistance to mutagenic stress.
Asunto(s)
Reparación del ADN , Elementos Transponibles de ADN , Proteínas de Unión al ADN , Genes Bacterianos , Hemiterpenos , Lactococcus lactis/genética , Lactococcus lactis/efectos de la radiación , Mutagénesis Insercional , Pentanos , Proteínas Bacterianas/genética , Butadienos/metabolismo , Pared Celular/metabolismo , Clonación Molecular , Análisis Mutacional de ADN , ADN Polimerasa I/metabolismo , Lactococcus lactis/efectos de los fármacos , Mitomicina/farmacología , Datos de Secuencia Molecular , Purinas/metabolismo , Pirimidinas/metabolismo , Recombinación Genética , Rayos UltravioletaRESUMEN
Microsequencing of a polypeptide with MW of 14.5 and pI of 5.0 induced by heat treatment at 42 degrees C and 50 degrees C in Lactococcus lactis subsp. lactis revealed that it corresponds to the co-chaperonin GroES. Quantitative analysis of analytical 2-D gels showed a relative induction of 12- and 11-fold after 30 min of heat adaptation at 42 degrees C and 50 degrees C, respectively. GroES is also induced by an acid shift from pH 7 to pH 5.5 and by UV254 nm-irradiation, with relative induction factors of 3.8 and 2.3, respectively. To our knowledge this is the first report showing induction of GroES by mild acid treatment. Contrasting to the relative induction of the groEL gene product, the second protein encoded by the groESL operon, GroES shows significantly higher induction under all stress situations.
Asunto(s)
Chaperonina 10/genética , Chaperonina 60/genética , Regulación Bacteriana de la Expresión Génica/fisiología , Lactococcus lactis/genética , Secuencia de Aminoácidos , Chaperonina 10/química , Chaperonina 60/química , Regulación Bacteriana de la Expresión Génica/efectos de la radiación , Calor , Concentración de Iones de Hidrógeno , Punto Isoeléctrico , Ácido Láctico/farmacología , Lactococcus lactis/efectos de la radiación , Datos de Secuencia Molecular , Peso Molecular , Análisis de Secuencia , Rayos UltravioletaRESUMEN
Rates of radiolytic inactivation of bacteria suspended in N2O-saturated solutions were dramatically increased over normal background levels when the media contained chloride or bicarbonate ions. The bacteria could be protected from this enhanced toxicity by the addition of free radical scavengers (ethanol, ascorbate, hydrogen peroxide, mannitol, glucose, EDTA, picolinic acid), indicating that the lethal reactions were extracellular in origin. Prior irradiation of chloride-containing solutions led to formation of hypochlorous acid, which was identified by detection of ring-chlorinated products when reacted with fluorescein. Prolonged irradiation of other solutions did not lead to accumulation of bactericidal agents; however, irradiation of bicarbonate-containing solutions in the presence of the spin trap 5,5-dimethyl-1-pyrroline N-oxide (DMPO) led to formation of the EPR-detectable DMPO.CO3- adduct. The results are interpreted in terms of formation of secondary radicals, among which the carbonate and chlorine radicals are uniquely toxic to bacteria. From rate comparisons of the solution components, it was concluded that the reactions involving chloride ion are unlikely to be expressed in biological environments, but that the CO3- radical could be an important intermediary oxidant in peroxide-inflicted cellular damage, particularly in spatially confined environments such as the leukocyte phagosome.
Asunto(s)
Antibacterianos/toxicidad , Escherichia coli/efectos de los fármacos , Radical Hidroxilo/toxicidad , Lactococcus lactis/efectos de los fármacos , Pseudomonas aeruginosa/efectos de los fármacos , Ácido Ascórbico/farmacología , Sinergismo Farmacológico , Ácido Edético/farmacología , Escherichia coli/crecimiento & desarrollo , Escherichia coli/efectos de la radiación , Etanol/farmacología , Depuradores de Radicales Libres , Rayos gamma , Glucosa/farmacología , Peróxido de Hidrógeno/farmacología , Cinética , Lactococcus lactis/crecimiento & desarrollo , Lactococcus lactis/efectos de la radiación , Luz , Manitol/farmacología , Modelos Biológicos , Ácidos Picolínicos/farmacología , Pseudomonas aeruginosa/crecimiento & desarrollo , Pseudomonas aeruginosa/efectos de la radiaciónRESUMEN
The temperate lactococcal phage TP901-1, induced by UV light from Lactococcus lactis subsp. cremoris 901-1, was characterized. The restriction map was found to be circular, and the packaging of TP901-1 DNA was concluded to occur by a headful mechanism. The pac region was localized on the 38.4-kb phage genome. TP901-1 belongs to the class of P335 phages (V. Braun, S. Hertwig, H. Neve, A. Geis, and M. Teuber, J. Gen. Microbiol. 135:2551-2560, 1989). Evidence is presented that the phages TP936-1 (V. Braun, S. Hertwig, H. Neve, A. Geis, and M. Teuber, J. Gen. Microbiol. 135:2551-2560, 1989) and C3-T1 (A. W. Jarvis, V. R. Parker, and M. B. Bianchin, Can. J. Microbiol. 38:398-404, 1992) are very closely related to or are identical to TP901-1. The lytically propagated TP901-1 phages were able to lysogenize both indicator strains Lactococcus cremoris 3107 and Wg2. Lysogenization resulted in site-specific integration of the phage genome into the bacterial chromosome. Only one chromosomal attB site was found in 20 independent lysogens. The attP region of TP901-1 and the attL and attR regions were cloned and sequenced. The results showed a core region of only 5 bp, in which the recombination occurs, followed after a 1-bp mismatch by a 7-bp identical region, TCAAT(T/C)AAGGTAA. This result was further verified by sequencing of the attB region obtained by PCR. An integration vector was constructed with the 6.5-kb EcoRI fragment from TP901-1 containing attP. This vector also functions in the plasmid-free strains, MG1363 and LM0230 with only one specific attB site, strongly indicating a more general use of the TP901-1-based integration vector in lactococci.
Asunto(s)
Bacteriófagos/genética , Vectores Genéticos/genética , Lactococcus lactis , Lisogenia/genética , Integración Viral/genética , Bacteriófagos/efectos de la radiación , Secuencia de Bases , Clonación Molecular , ADN Circular/genética , Genes Bacterianos/genética , Lactococcus lactis/efectos de la radiación , Datos de Secuencia Molecular , Recombinación Genética , Mapeo Restrictivo , Rayos UltravioletaRESUMEN
The growth of R-, S- and M (g)-dissociants of Streptococcus lactis, Bacillus coagulans, Rhodococcus rubropertinctus under the action of some physico-chemical factors: temperature, pH, ultraviolet (UV) rays, high concentration of NaCl and storage have been compared. R-variants gain selective advantage under the influence of UV-irradiation, high temperature and storage; S-variants--at decreasing of active pH of medium; M (g)-variants--at decreasing of growth temperature, high values of pH, increased NaCl concentration. The dissociation has been concluded to enlarge the limits of the species survival.
Asunto(s)
Bacterias Grampositivas/crecimiento & desarrollo , Bacillus/efectos de los fármacos , Bacillus/crecimiento & desarrollo , Bacillus/efectos de la radiación , Fenómenos Químicos , Química Física , Bacterias Grampositivas/efectos de los fármacos , Bacterias Grampositivas/efectos de la radiación , Concentración de Iones de Hidrógeno , Lactococcus lactis/efectos de los fármacos , Lactococcus lactis/crecimiento & desarrollo , Lactococcus lactis/efectos de la radiación , Preservación Biológica , Rhodococcus/efectos de los fármacos , Rhodococcus/crecimiento & desarrollo , Rhodococcus/efectos de la radiación , Cloruro de Sodio/farmacología , Temperatura , Rayos UltravioletaRESUMEN
We have identified in Lactococcus lactis, an analogue of Escherichia coli RecA protein. Physiological responses such as ultraviolet (UV) and chemical mutagenesis and induction of prophage have been characterized and suggest the existence of RecA-like functions in this commercially important species. The putative RecA protein was detected at the position of an apparent molecular weight of 39 kDa by Western blot analysis by using antiserum against E coli RecA protein. In addition, the protein level is significantly increased after UV irradiation in a wild-type strain compared to the recombination deficient mutant strain.
Asunto(s)
Lactococcus lactis/análisis , Rec A Recombinasas/análisis , Western Blotting , Lactococcus lactis/metabolismo , Lactococcus lactis/efectos de la radiación , Peso Molecular , Mutación , Rec A Recombinasas/química , Rec A Recombinasas/metabolismo , Recombinación Genética , Rayos UltravioletaRESUMEN
The lethal and mutagenic effects of various mutagens on three strains of Streptococcus lactis were investigated. Lethality studies demonstrated that S. lactis was relatively sensitive to UV irradiation, methyl methanesulphonate (MMS) and N-methyl-N'-nitro-N-nitrosoguanidine (MNNG), and, to a lesser extent, to ethyl methanesulphonate (EMS). A spontaneous derivative Lac-, which has lost a 37-Md plasmid, was slightly more resistant and much less mutable than the wild-type after UV irradiation. Although the three strains were strongly mutated by EMS for the genetic marker assayed (Rifr), an increase in the mutation frequency was also observed after MMS and MNNG treatments.
Asunto(s)
Alquilantes/toxicidad , Lactococcus lactis/genética , Mutación , Rayos Ultravioleta/efectos adversos , ADN Bacteriano/análisis , Electroforesis en Gel de Agar , Lactococcus lactis/efectos de los fármacos , Lactococcus lactis/efectos de la radiación , PlásmidosRESUMEN
Plasmid pIL7 (33 kilobases) from Streptococcus lactis enhances UV resistance and prophage stability. A 5.4-kilobase pIL7 fragment carrying genes coding for both characters was cloned into S. lactis, using plasmid pHV1301 as the cloning vector. The recombinant plasmid was subsequently transferred to three other S. lactis strains by transformation or protoplast fusion. Cloned genes were expressed in all tested strains.
Asunto(s)
Bacteriófagos/genética , Clonación Molecular , Lactococcus lactis/efectos de la radiación , Plásmidos , Rayos Ultravioleta , Bacillus subtilis/genética , Bacteriófagos/fisiología , Enzimas de Restricción del ADN , Vectores Genéticos , Genotipo , Lactococcus lactis/genética , FenotipoRESUMEN
A recombination-deficient mutant of Streptococcus lactis ML3 designated MMS36 was isolated on the basis of its sensitivity to methyl methanesulfonate. This mutant also displayed sensitivity to UV irradiation. The inability of MMS36 to mediate homologous recombination was demonstrated by transduction of plasmid-linked lactose fermenting ability but not chromosomally mediated streptomycin resistance.