RESUMEN
AIMS: (i) To determine the aflatoxin B1 (AFB1 ) adsorption and desorption dynamics in the presence of Lactobacillus rhamnosus RC007 under simulated transit of AFB1 at each gastrointestinal tract (GIT-saliva, stomach and intestine) stage consecutively and then, separately, (ii) to study the ability of L. rhamnosus RC007 to biotransform AFB1 as a strategy that complements the adsorption process. METHODS AND RESULTS: The AFB1 adsorption and desorption assay simulating the GIT passage of AFB1 (93·89 ng g-1 ) in the presence of L. rhamnosus RC007 (108 CFU per ml) was conducted. Moreover, lactic acid production was determined. Results demonstrated that predominant environmental conditions in salivary solution induced a low AFB1 adsorption, while the transit through the gastric solution and intestinal solution allowed high percentages of adsorption and did not generate significant AFB1 desorption. CONCLUSIONS: The AFB1 adsorption and desorption dynamics in the presence of L. rhamnosus RC007 was favoured by gastric and intestinal environment. SIGNIFICANCE AND IMPACT OF THE STUDY: The knowledge of the adsorption dynamics of AFB1 with a micro-organism of interest will allow predicting its behaviour at each stage of the GIT.
Asunto(s)
Aflatoxina B1/metabolismo , Tracto Gastrointestinal/metabolismo , Lacticaseibacillus rhamnosus/metabolismo , Adsorción , Animales , Tracto Gastrointestinal/microbiología , Ácido Láctico/metabolismo , Modelos BiológicosRESUMEN
AIMS: To investigate multiple tolerance of Saccharomyces cerevisiae obtained through a laboratory strategy of adaptive evolution in acetic acid, its relation with enzymatic ROS detoxification and bioethanol 2G production. METHODS AND RESULTS: After adaptive evolution in acetic acid, a clone (Y8A) was selected for its tolerance to high acetic acid concentrations (13 g l-1 ) in batch cultures. Y8A was resistant to multiple stresses: osmotic, thermic, oxidative, saline, ethanol, organic acid, phenolic compounds and slow freeze-thawing cycles. Also, Y8A was able to maintain redox homeostasis under oxidative stress, whereas the isogenic parental strain (Y8) could not, indicating higher basal activity levels of antioxidative enzyme Catalase (CAT) and Gluthatione S-transferase (GST) in Y8A. Y8A reached higher bioethanol levels in a fermentation medium containing up to 8 g l-1 of acetic acid when compared to parental strain Y8. CONCLUSIONS: A multiple-stress-tolerant clone was obtained using adaptive evolution in acetic acid. Stress cross-tolerance could be explained by its enzymatic antioxidative capacity, namely CAT and GST. SIGNIFICANCE AND IMPACT OF THE STUDY: We demonstrate that adaptive evolution used in S. cerevisiae was a useful strategy to obtain a yeast clone tolerant to multiple stresses. At the same time, our findings support the idea that tolerance to oxidative stress is the common basis for stress cotolerance, which is related to an increase in the specific enzymes CAT and GST but not in Superoxide dismutase, emphasizing the fact that detoxification of H2 O2 and not O2 Ë is a key condition for multiple stress tolerance in S. cerevisiae.
Asunto(s)
Ácido Acético/farmacología , Antioxidantes/metabolismo , Etanol/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae , Saccharomyces cerevisiae/efectos de los fármacos , Saccharomyces cerevisiae/enzimología , Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/fisiologíaRESUMEN
AIM: To evaluate the ability of probiotic Saccharomyces cerevisiae RC016 strain to reduce fumonisin B(1) (FB(1)) in vitro and to optimize the culture conditions for the growth of the yeast employing surface response methodology. METHODS AND RESULTS: Using Plackett-Burman screening designs (PBSD) and central composite designs (CCD), an optimized culture medium containing (g l(-1)) fermentable sugars provided by sugar cane molasses (CMs), yeast extract (YE) and (NH(4))(2) HPO(4) (DAP) was formulated. The S. cerevisiae RC016 strain showed the greatest binding at all assayed FB1 concentration. The CMs, YE, DAP concentrations and incubation time influenced significantly the biomass of S. cerevisiae RC016. CONCLUSION: A combination of CMs 17%; YE 4·61 g l(-1) and incubation time 60 h was optimum for maximum biomass of S. cerevisiae RC016. SIGNIFICANCE AND IMPACT OF THE STUDY: The importance of this work lies in the search for live strains with both probiotic and fumonisin B1 decontamination properties that could be sustainably produced in a medium just containing cheap carbon, nitrogen and phosphorus sources and would be included in a novel product to animal feed.
Asunto(s)
Biomasa , Fumonisinas/química , Probióticos , Saccharomyces cerevisiae/metabolismo , Alimentación Animal , Reactores Biológicos , Carbono/metabolismo , Medios de Cultivo/química , Fermentación , Microbiología Industrial , Modelos Estadísticos , Melaza , Nitrógeno/metabolismo , Saccharomyces cerevisiae/crecimiento & desarrollo , SaccharumRESUMEN
AIM: To maximize biomass production of an ochratoxigenic mould-controlling strain of Lachancea thermotolerans employing response surface methodology (RSM). METHODS AND RESULTS: Using Plackett-Burman screening designs (PBSD) and central composite designs (CCD), an optimized culture medium containing (g l(-1) ): fermentable sugars (FS), 139·2, provided by sugar cane molasses (CMz), (NH(4) )(2) HPO(4) (DAP), 9·0, and yeast extract (YE), 2·5, was formulated. Maximal cell concentration obtained after 24 h at 28°C was 24·2 g l(-1) cell dry weight (CDW). The mathematical model obtained was validated in experiments performed in shaken-flask cultures and also in aerated bioreactors. Maximum yield and productivity values achieved were, respectively, of 0·23 g CDW/g FS in a medium containing (g l(-1) ): FS, 87·0; DAP, 7·0; YE, 1·0; and of 0·96 g CDW l(-1) h(-1) in a medium containing (g l(-1) ): FS, 150·8 plus DAP, 6·9. CONCLUSIONS: Optimized culture conditions for maximizing yeast biomass production determined in flask cultures were applicable at a larger scale. The highest yield values were attained in media containing relatively low-CMz concentrations supplemented with DAP and YE. Yeast extract would not be necessary if higher productivity is the aim. SIGNIFICANCE AND IMPACT OF THE STUDY: Cells of L. thermotolerans produced aerobically could be sustainably produced in a medium just containing cheap carbon, nitrogen and phosphorus sources. Response surface methodology allowed the fine-tuning of cultural conditions.
Asunto(s)
Agentes de Control Biológico , Hongos/crecimiento & desarrollo , Levaduras/crecimiento & desarrollo , Antibiosis , Biomasa , Medios de Cultivo , Fermentación , Estadística como Asunto , Levaduras/fisiologíaRESUMEN
AIMS: Analysis of the physiology and metabolism of Escherichia coli arcA and creC mutants expressing a bifunctional alcohol-acetaldehyde dehydrogenase from Leuconostoc mesenteroides growing on glycerol under oxygen-restricted conditions. The effect of an ldhA mutation and different growth medium modifications was also assessed. METHODS AND RESULTS: Expression of adhE in E. coli CT1061 [arcA creC(Con)] resulted in a 1.4-fold enhancement in ethanol synthesis. Significant amounts of lactate were produced during micro-oxic cultures and strain CT1061LE, in which fermentative lactate dehydrogenase was deleted, produced up to 6.5 +/- 0.3 g l(-1) ethanol in 48 h. Escherichia coli CT1061LE derivatives resistant to >25 g l(-1) ethanol were obtained by metabolic evolution. Pyruvate and acetaldehyde addition significantly increased both biomass and ethanol concentrations, probably by overcoming acetyl-coenzyme A (CoA) shortage. Yeast extract also promoted growth and ethanol synthesis, and this positive effect was mainly attributable to its vitamin content. Two-stage bioreactor cultures were conducted in a minimal medium containing 100 microg l(-1) calcium d-pantothenate to evaluate oxic acetyl-CoA synthesis followed by a switch into fermentative conditions. Ethanol reached 15.4 +/- 0.9 g l(-1) with a volumetric productivity of 0.34 +/- 0.02 g l(-1) h(-1). CONCLUSIONS: Escherichia coli responded to adhE over-expression by funnelling carbon and reducing equivalents into a highly reduced metabolite, ethanol. Acetyl-CoA played a key role in micro-oxic ethanol synthesis and growth. SIGNIFICANCE AND IMPACT OF THE STUDY: Insight into the micro-oxic metabolism of E. coli growing on glycerol is essential for the development of efficient industrial processes for reduced biochemicals production from this substrate, with special relevance to biofuels synthesis.
Asunto(s)
Alcohol Deshidrogenasa/metabolismo , Aldehído Oxidorreductasas/metabolismo , Escherichia coli/genética , Etanol/metabolismo , Glicerol/metabolismo , Leuconostoc/enzimología , Acetilcoenzima A/metabolismo , Alcohol Deshidrogenasa/genética , Aldehído Oxidorreductasas/genética , Escherichia coli/metabolismo , Mutación , Oxidación-ReducciónRESUMEN
AIMS: To study the effect of prestress conditions on the freezing and thawing (FT) response of two baker's yeast strains and the use of statistical analysis to optimize resistance to freezing. METHODS AND RESULTS: Tolerance to FT of industrial strains of Saccharomyces cerevisiae was associated to their osmosensitivity and growth phase. Pretreatments with sublethal stresses [40 degrees C, 0.5 mol l(-1) NaCl, 1.0 mol l(-1) sorbitol or 5% (v/v) ethanol] increased freeze tolerance. Temperature or hyperosmotic prestresses increased trehalose contents, nevertheless no clear correlation was found with improved FT tolerance. Plackett-Burman design and response surface methodology were applied to improve freeze tolerance of the more osmotolerant strain. Optimal prestress conditions found were: 0.779 mol l(-1) NaCl, 0.693% (v/v) ethanol and 32.15 degrees C. CONCLUSIONS: Ethanol, saline, osmotic or heat prestresses increased freezing tolerance of two phenotypically distinct baker's yeast strains. A relationship among prestresses, survival and trehalose content was not clear. It was possible to statistically find optimal combined prestress conditions to increase FT tolerance of the osmotolerant strain. SIGNIFICANCE AND IMPACT OF THE STUDY: Statistically designed combination of prestress conditions that can be applied during the production of baker's yeast could represent a useful tool to increase baker's yeast FT resistance.
Asunto(s)
Microbiología de Alimentos , Modelos Estadísticos , Saccharomyces cerevisiae/fisiología , Etanol/análisis , Congelación , Glucosa/análisis , Viabilidad Microbiana , Modelos Biológicos , Saccharomyces cerevisiae/metabolismo , Trehalosa/análisisRESUMEN
Different natural antimicrobials affected viability of bacterial contaminants isolated at critical steps during a beer production process. In the presence of 1 mg/ml chitosan and 0.3 mg/ml hops, the viability of Escherichia coli in an all malt barley extract wort could be reduced to 0.7 and 0.1% respectively after 2 hour- incubation at 4 degrees C. The addition of 0.0002 mg/ml nisin, 0.1 mg/ml chitosan or 0.3 mg/ml hops, selectively inhibited growth of Pediococcus sp. in more than 10,000 times with respect to brewing yeast in a mixed culture. In the presence of 0.1 mg ml chitosan in beer, no viable cells of the thermoresistant strain Bacillus megaterium were detected. Nisin, chitosan and hops increased microbiological stability during storage of a local commercial beer inoculated with Lactobacillus plantarum or Pediococcus sp. isolated from wort. Pulsed Electric Field (PEF) (8 kV/cm, 3 pulses) application enhanced antibacterial activity of nisin and hops but not that of chitosan. The results herein obtained suggest that the use of these antimicrobial compounds in isolation or in combination with PEF would be effective to control bacterial contamination during beer production and storage.
Asunto(s)
Bacillus megaterium/aislamiento & purificación , Cerveza/microbiología , Quitosano/farmacología , Campos Electromagnéticos , Escherichia coli/aislamiento & purificación , Humulus , Microbiología Industrial/métodos , Lactobacillus plantarum/aislamiento & purificación , Nisina/farmacología , Pediococcus/aislamiento & purificación , Extractos Vegetales/farmacología , Bacillus megaterium/efectos de los fármacos , Bacillus megaterium/crecimiento & desarrollo , Bacillus megaterium/efectos de la radiación , Escherichia coli/efectos de los fármacos , Escherichia coli/crecimiento & desarrollo , Escherichia coli/efectos de la radiación , Fermentación , Conservación de Alimentos , Lactobacillus plantarum/efectos de los fármacos , Lactobacillus plantarum/crecimiento & desarrollo , Lactobacillus plantarum/efectos de la radiación , Pruebas de Sensibilidad Microbiana , Pediococcus/efectos de los fármacos , Pediococcus/crecimiento & desarrollo , Pediococcus/efectos de la radiación , TemperaturaRESUMEN
Different natural antimicrobials affected viability of bacterial contaminants isolated at critical steps during a beer production process. In the presence of 1 mg/ml chitosan and 0.3 mg/ml hops, the viability of Escherichia coli in an all malt barley extract wort could be reduced to 0.7 and 0.1% respectively after 2 hour- incubation at 4 °C. The addition of 0.0002 mg/ml nisin, 0.1 mg/ml chitosan or 0.3 mg/ml hops, selectively inhibited growth of Pediococcus sp. in more than 10,000 times with respect to brewing yeast in a mixed culture. In the presence of 0.1mg ml chitosan in beer, no viable cells of the thermoresistant strain Bacillus megaterium were detected. Nisin, chitosan and hops increased microbiological stability during storage of a local commercial beer inoculated with Lactobacillus plantarum or Pediococcus sp. isolated from wort. Pulsed Electric Field (PEF) (8 kV/cm, 3 pulses) application enhanced antibacterial activity of nisin and hops but not that of chitosan. The results herein obtained suggest that the use of these antimicrobial compounds in isolation or in combination with PEF would be effective to control bacterial contamination during beer production and storage.
Diferentes antimicrobianos naturales disminuyeron la viabilidad de bacterias contaminantes aisladas en etapas críticas del proceso de producción de cerveza. En un extracto de malta, el agregado de 1 mg/ml de quitosano y de 0,3 mg ml de lúpulo permitió reducir la viabilidad de Escherichia coli a 0,7 y 0,1%, respectivamente, al cabo de 2 horas de incubación a 4 °C. El agregado de 0,0002 mg/ml de nisina, 0,1 mg/ml de quitosano o de 0,3 mg/ml de lúpulo inhibió selectivamente (10.000 veces más) el crecimiento de Pediococcus sp. respecto de la levadura de cerveza en un cultivo mixto. El agregado de 0,1 mg/ml de quitosano permitió disminuir la viabilidad de una cepa bacteriana termorresistente, Bacillus megaterium, hasta niveles no detectables. Por otra parte, el agregado de nisina, quitosano y lúpulo aumentó la estabilidad microbiológica durante el almacenamiento de cervezas inoculadas con Lactobacillus plantarum y Pediococcus sp. aislados de mosto de cerveza. La aplicación de campos eléctricos pulsantes (CEP) (3 pulsos de 8kV/cm) aumentó el efecto antimicrobiano de la nisina y del lúpulo, pero no el del quitosano. Los resultados obtenidos indicarían que el uso de antimicrobianos naturales en forma individual o en combinación con CEP puede constituir un procedimiento efectivo para el control de la contaminación bacteriana durante el proceso de elaboración y almacenamiento de la cerveza.
Asunto(s)
Bacillus megaterium/aislamiento & purificación , Cerveza/microbiología , Quitosano/farmacología , Campos Electromagnéticos , Escherichia coli/aislamiento & purificación , Humulus , Microbiología Industrial/métodos , Lactobacillus plantarum/aislamiento & purificación , Nisina/farmacología , Pediococcus/aislamiento & purificación , Extractos Vegetales/farmacología , Bacillus megaterium/efectos de los fármacos , Bacillus megaterium/crecimiento & desarrollo , Bacillus megaterium/efectos de la radiación , Escherichia coli/efectos de los fármacos , Escherichia coli/crecimiento & desarrollo , Escherichia coli/efectos de la radiación , Fermentación , Conservación de Alimentos , Lactobacillus plantarum/efectos de los fármacos , Lactobacillus plantarum/crecimiento & desarrollo , Lactobacillus plantarum/efectos de la radiación , Pruebas de Sensibilidad Microbiana , Pediococcus/efectos de los fármacos , Pediococcus/crecimiento & desarrollo , Pediococcus/efectos de la radiación , TemperaturaRESUMEN
Rising activity determined as CO2 production of two commercial strains of Saccharomyces cerevisiae could be increased mainly in sweet bread doughs by introducing a "starvation/pulse feeding" schedule of sugar cane molasses during a fed-batch propagation. Such increase was strain dependent. Except for the trehalose intracellular level, other traits related to the yeast industrial performance were unaffected. Applicability of method for baker's yeast industrial production is discussed.
Asunto(s)
Dióxido de Carbono/metabolismo , Microbiología Industrial/métodos , Micología/métodos , Saccharomyces cerevisiae/crecimiento & desarrollo , Pan , Metabolismo de los Hidratos de Carbono , Dióxido de Carbono/análisis , Fermentación , Melaza , Saccharomyces cerevisiae/metabolismo , Trehalosa/metabolismoRESUMEN
La actividad panificante valorada como producción de CO2 de dos cepas comerciales de Saccharomyces cerevisiae pudo ser incrementada, principalmente en amasijos azucarados, por la aplicación de un esquema de hambreado/ pulso¼ de melaza de caña de azúcar durante su propagación bajo la forma de lote alimentado. Dicho incremento fue dependiente de la cepa utilizada. Otras características relacionadas con el comportamiento industrial de las levaduras no se vieron afectadas, con excepción de la concentración intracelular de trehalosa.Se discute la aplicabilidad del método para la producción industrial de levaduras de panificación.
Baking activity determined as CO2 production of two commercial strains of Saccharomyces cerevisiae could be increased mainly in sweet bread doughs by introducing a starvation/ pulse feeding schedule of sugar cane molasses during a fed-batch propagation . Such increase was strain dependent. Except for the trehalose intracellular level, other traits related to the yeast industrial performance were unaffected. Applicability of method for bakers yeast industrial production is discussed.
Asunto(s)
Dióxido de Carbono/metabolismo , Microbiología Industrial/métodos , Micología/métodos , Saccharomyces cerevisiae/crecimiento & desarrollo , Pan , Metabolismo de los Hidratos de Carbono , Dióxido de Carbono/análisis , Fermentación , Melaza , Saccharomyces cerevisiae/metabolismo , Trehalosa/metabolismoRESUMEN
Rising activity determined as CO2 production of two commercial strains of Saccharomyces cerevisiae could be increased mainly in sweet bread doughs by introducing a [quot ]starvation/pulse feeding[quot ] schedule of sugar cane molasses during a fed-batch propagation. Such increase was strain dependent. Except for the trehalose intracellular level, other traits related to the yeast industrial performance were unaffected. Applicability of method for bakers yeast industrial production is discussed.
RESUMEN
The effects of vacuum-drying and freeze-drying on the cell viability of a commercial baker's yeast, Saccharomyces cerevisiae, strain with different endogenous contents of trehalose were analyzed. An osmotolerant Zygosaccharomyces rouxii strain was used for comparative purposes. Higher viability values were observed in cells after vacuum-drying than after freeze-drying. Internal concentrations of trehalose in the range 10-20% protected cells in both dehydration processes. Endogenous trehalose concentrations did not affect the water sorption isotherm nor the Tg values. The effect of external matrices of trehalose and maltodextrin was also studied. The addition of external trehalose improved the survival of S. cerevisiae cells containing 5% internal trehalose during dehydration. Maltodextrin (1.8 kDa) failed to protect vacuum-dried samples at 40 degrees C. The major reduction in the viability during the freeze-drying process of the sensitive yeast cells studied was attributed to the freezing step. The suggested protective mechanisms for each particular system are vitrification and the specific interactions of trehalose with membranes and/or proteins. The failure of maltodextrins to protect cells was attributed to the fact that none of the suggested mechanisms of protection could operate in these systems.
Asunto(s)
Saccharomyces cerevisiae/fisiología , Trehalosa/análisis , Liofilización , Saccharomyces cerevisiae/químicaRESUMEN
Differential scanning calorimetry thermograms of various samples of commercial instant active dry yeasts revealed a clear glass transition typical of amorphous carbohydrates and sugars. The resulting glass transition temperatures were found to decrease with increasing moisture content. The observed glass curve was similar to that of pure trehalose, which is known to accumulate in large amounts in baker's yeast. The effect of heat treatment at various temperatures on the fermentative activity (as measured by the metabolic production of CO(2)) of dry yeast was studied. First-order plots were obtained representing the loss of fermentative activity as a function of heating time at the various temperatures assayed. Significant losses of fermentative activity were observed in vitrified yeast samples. The dependence of rate constants with temperature was found to follow Arrhenius behavior. The relationship between the loss of fermentative activity and glass transition was not verified, and the glass transition was not reflected on the temperature dependence of fermentative activity loss.
Asunto(s)
Calor , Levaduras/metabolismo , Biotecnología/métodos , Rastreo Diferencial de Calorimetría , Dióxido de Carbono/metabolismo , Fermentación , Temperatura , Levaduras/química , Levaduras/fisiologíaRESUMEN
Incubation of Candida albicans yeast cells with human luteinizing hormone (hLH), human chorionic gonadotrophin (hCG) or glucagon produced a significant rise in cAMP total levels. The effect of these hormones in permeabilized cells of the fungus produced a 2-3 fold increase in the Mg2+, GTP-dependent adenylyl cyclase activity as well as full activation of the cAMP-dependent protein kinase (PKA) activity. These results indicate that the interaction of the mammalian hormones with the fungus triggered the cAMP activation cascade in a similar way to that found in higher eukaryotic organisms.
Asunto(s)
Candida albicans/metabolismo , Gonadotropina Coriónica/farmacología , AMP Cíclico/metabolismo , Glucagón/farmacología , Hormona Luteinizante/farmacología , Adenilil Ciclasas/metabolismo , Permeabilidad de la Membrana Celular/efectos de los fármacos , AMP Cíclico/análisis , AMP Cíclico/biosíntesis , Humanos , Magnesio/análisis , Magnesio/metabolismo , Proteínas Quinasas/metabolismo , Sistemas de Mensajero Secundario , Tolueno/farmacologíaRESUMEN
A cAMP-dependent protein kinase from mycelia of Saccobolus platensis was characterized. The holoenzyme seems to be a dimer (i.e., regulatory subunit--catalytic subunit) of 78,000 Da, slightly activated by cAMP but susceptible to dissociation into its subunits by cAMP, or by kemptide and protamine, the best substrates for Saccobolus protein kinase. The regulatory subunit was purified to homogeneity by affinity chromatography. It is highly specific for cAMP and has two types of binding sites but failed to inhibit the phosphotransferase activity of the homologous or the heterologous (bovine heart) catalytic components. The activity of the catalytic subunit was completely abolished by the regulatory component of the bovine heart protein kinase as well as by a synthetic peptide corresponding to the active site of the mammalian protein kinase inhibitor. The data suggest that interaction between the subunits of the S. platensis protein kinase is different than that found in cAMP-dependent protein kinases from other sources. Similarities and differences between the Saccobolus protein kinase and enzymes from low eucaryotes and mammalian tissues are discussed.
Asunto(s)
Ascomicetos/enzimología , Proteínas Quinasas/aislamiento & purificación , Centrifugación por Gradiente de Densidad , Cromatografía de Afinidad , Cromatografía DEAE-Celulosa , AMP Cíclico/metabolismo , Citosol/enzimología , Electroforesis en Gel de Poliacrilamida , Cinética , Sustancias Macromoleculares , Peso Molecular , Conformación Proteica , Proteínas Quinasas/metabolismo , Especificidad por SustratoRESUMEN
GTP, GTP-gamma-S and Gpp(NH)p produced a significant activation of Mg2(+)-dependent adenylyl cyclase in permeabilized cells of Candida albicans. This activation was inhibited by GDP-beta-S. Maximal stimulation (4-6 fold) was obtained when Mg2+ and GTP-gamma-S were added during permeabilization. The guanine nucleotide-stimulated activity could be further activated by glucagon in a dose dependent manner being the maximal stimulation (4-5 fold) attained at 10(-6) M glucagon. Addition of the hormone to yeast cells incubated under conditions conducive to produce germ tubes blocked hyphal development and promoted multibudded cell morphology.
Asunto(s)
Adenilil Ciclasas/metabolismo , Candida albicans/enzimología , Glucagón/farmacología , Guanosina Trifosfato/análogos & derivados , Guanosina Trifosfato/farmacología , Guanilil Imidodifosfato/farmacología , Tionucleótidos/farmacología , Candida albicans/citología , Candida albicans/efectos de los fármacos , Activación Enzimática , Guanosina 5'-O-(3-Tiotrifosfato) , Cinética , Magnesio/farmacologíaRESUMEN
Intracellular levels of cAMP and specific activities of adenylate cyclase, cAMP phosphodiesterase and cAMP-dependent protein kinase were measured during filamentation in the dimorphic fungus Candida albicans. Enzymatic assays were performed in permeabilized cells under conditions prevented endogenous proteolysis. The variations observed in cAMP levels were mainly accounted for by variations in the specific activities of adenylate cyclase and cAMP phosphodiesterase at different stages during germ tube formation. cAMP-dependent protein kinase, measured with kemptide as exogenous substrate, was developmental regulated. Some properties of the enzymatic activities from cell-free extracts are described.