Dolores de Arriaga - Academia.edu (original) (raw)

Papers by Dolores de Arriaga

Journal of Dairy Science, Aug 1, 2013

PubMed, May 1, 1991

High concentrations of either Mg-ATP complex, free ATP, or free Mg2+ ions were inhibitors of the ... more High concentrations of either Mg-ATP complex, free ATP, or free Mg2+ ions were inhibitors of the mitochondrial F1-ATPase moiety from Phycomyces blakesleeanus. Free Mg2+ acts as a linear competitive inhibitor with regard to Mg-ATP hydrolysis with a Ki value of 2.8 mM. The inhibition by free ATP was markedly biphasic and thus simple competitive inhibition alone is not sufficient to explain the inhibitory effect. From these results conclusions were drawn about the binding of the substrate, Mg-ATP complex, to the enzyme.

PubMed, Aug 1, 1992

Exposure of lactate dehydrogenase from rabbit muscle to the Fe(III)/EDTA/ascorbate oxidation syst... more Exposure of lactate dehydrogenase from rabbit muscle to the Fe(III)/EDTA/ascorbate oxidation system leads to a time-dependent enzymatic inactivation (rate of inactivation of 7.35 x 10(-3) min-1), as well as to a spontaneous fragmentation of the protein. Fe(III) is the most important compound in this system, having the highest inactivating effects at concentrations above 10 microM. The substrate pyruvate and the products of the enzymatic reaction, when added at high concentration to the full mixture of the system, have a partial protective effect on the catalytic activity.

PubMed, Mar 1, 1989

The inductive effect of different sugars on beta-galactosidase synthesis in Phycomyces blakesleea... more The inductive effect of different sugars on beta-galactosidase synthesis in Phycomyces blakesleeanus has been studied. The enzyme was inducible by galactose and fructose. When grown on these sugars the enzyme level was 10-20 times greater than when grown on glucose. We have detected both intra- and extracellular beta-galactosidase activity when Phycomyces blakesleeanus was grown on galactose, but only extracellular beta-galactosidase activity when grown on fructose plus lactose.

Journal of Bacteriology, Nov 1, 1989

Food Control, Aug 1, 2012

ABSTRACT MIC values of seven pure phenolic compounds (thymol, carvacrol, eugenol, hydroquinone, p... more ABSTRACT MIC values of seven pure phenolic compounds (thymol, carvacrol, eugenol, hydroquinone, p-hydroxybenzoic acid, protocatechuic acid and gallic acid) were assessed against several strains of two Gram-positive (Staphylococcus aureus and Bacillus cereus) and two Gram-negative (Escherichia coli – including Shiga toxin-producing serogroups, STEC – and Pseudomonas fluorescens) bacteria, by using a standardized microdilution assay (ISO 20776-1:2006). Carvacrol and thymol were the most effective compounds for all genera/species studied, with the exception of S. aureus, for which hydroquinone was the most effective compound. However, we detected significant differences (p < 0.05) in the inhibition pattern of the four genera/species for both compounds (thymol and carvacrol), as well as for hydroquinone. Concerning to the less effective compounds, the four genera/species showed great differences among each other, hydroquinone being the least effective for E. coli, gallic acid for B. cereus and P. fluorescens, and eugenol for S. aureus. Gram-positive bacteria were more sensitive than Gram-negative ones for the majority of the compounds, although a general inter- and intraspecific pattern of behaviour in Gram-positive and Gram-negative bacteria was not detected. Each individual phenolic compound did not show a uniform antimicrobial effect on all strains included in each of the genera/species. Gallic acid and hydroquinone exhibited the highest antioxidant capacity and thymol and carvacrol the lowest. Gallic acid was effective in the control of S. aureus at lower concentrations than those used in the food industry as flavouring, and also showed important antioxidant ability. Therefore, this compound would be of interest in the control of S. aureus in foods.

Biochemistry and Cell Biology, Jul 1, 1991

Mitochondrial F1-ATPase was purified from the mycelium of Phycomyces blakesleeanus NRRL 1555(−) a... more Mitochondrial F1-ATPase was purified from the mycelium of Phycomyces blakesleeanus NRRL 1555(−) and its kinetic characteristics were studied. Sodium dodecyl sulfate – polyacrylamide gel electrophoresis of the enzyme reveals five bands (α, β, γ, δ, and ε) characteristic of the F1 portion with apparent molecular weights of 60 000, 53 000, 31 000, 25 000, and 21 000, respectively. The molecular weight of the native F1-ATPase from Phycomyces blakesleeanus was in agreement with the stoichiometry α3 β3 γ δ ε. The MgATP complex is the true substrate for ATPase activity which has a Km value of 0.15 mM. High concentrations of free ATP or free Mg2+ ions inhibit the ATPase activity. ADP appears to act as a negative allosteric effector with regard to MgATP hydrolysis, with the apparent Vmax remaining unchanged.Key words: mitochondrial F1-ATPase, Phycomyces blakesleeanus, enzyme purification, kinetic properties.

Journal of General Microbiology, Feb 1, 1993

The accumulation and mobilization of glycogen have been measured in Phycomyces blukesleeunus subj... more The accumulation and mobilization of glycogen have been measured in Phycomyces blukesleeunus subjected to a variety of environmental and nutritional conditions. Depending on the conditions and the stage in the life cycle, glycogen represented from 10 % to less than 1 YO of the dry weight of the mycelia. Maximum values were observed during the exponential growth phase irrespective of the carbon source used. The highest glycogen levels were detected with 2% (w/v) sorbitol as carbon source. A low glycogen content was present during growth on acetate. In these cultures glycogen was presumably synthesized from glyoxylate-dependent metabolites. Glycogen accumulated in cultures deprived specifically of nitrogen or phosphorus in the presence of an excess of a usable carbon source, suggesting that reserve carbohydrate accumulation is a general response to nutrient limitation. However, temporary starvation caused by removal of glucose or by transfer to media with relatively poor carbon sources such as acetate led to mobilization of glycogen in the mycelia. During sporulation the disappearance of glycogen was almost complete.

European Food Research and Technology, Dec 26, 2016

Applied and Environmental Microbiology, 1995

NADP-isocitrate dehydrogenase [isocitrate:NADP(sup+) oxidoreductase (decarboxylating); EC 1.1.1.4... more NADP-isocitrate dehydrogenase [isocitrate:NADP(sup+) oxidoreductase (decarboxylating); EC 1.1.1.42] was purified from Cephalosporium acremonium as a single species. The enzyme is a dimer of 140 kDa with identical subunits of 75 kDa. The existence of a monomer-dimer equilibrium is apparent as revealed by an enzyme dilution approach. The chelate complex of the tribasic form of isocitrate and Mg(sup2+) is the true substrate. The V(infmax) depends on a basic form of an ionizable group of the enzyme-substrate complex with a pK(infes) (pK of the enzyme-substrate complex) of 6.9 and a (Delta)H(infion) (activation enthalpy) of -2 (plusmn) 0.4 kcal mol(sup-1) (ca. 8 (plusmn) 2 kJ mol(sup-1)). The enzyme showed maximum activity at 60(deg)C, an unusually high temperature for a nonthermophilic fungus. The thermodynamic parameters for isocitrate oxidative decarboxylation and for the binding of isocitrate and NADP(sup+) were calculated. We analyzed the kinetic thermal stability of the enzyme at p...

Biochemical Journal, May 1, 1982

Fungal Genetics and Biology, Apr 1, 2002

Phycomyces blakesleeanus isocitrate lyase (EC 4.1.3.1) is in vivo reversibly inactivated by hydro... more Phycomyces blakesleeanus isocitrate lyase (EC 4.1.3.1) is in vivo reversibly inactivated by hydrogen peroxide. The purified enzyme showed reversible inactivation by an ascorbate plus Fe(2+) system under aerobic conditions. Inactivation requires hydrogen peroxide; was prevented by catalase, EDTA, Mg(2+), isocitrate, GSH, DTT, or cysteine; and was reversed by thiols. The ascorbate served as a source of hydrogen peroxide and also reduced the Fe(3+) ions produced in a &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;quot;site-specific&amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;quot; Fenton reaction. Two redox-active cysteine residues per enzyme subunit are targets of oxidative modification; one of them is located at the catalytic site and the other at the metal regulatory site. The oxidized enzyme showed covalent and conformational changes that led to inactivation, decreased thermal stability, and also increased inactivation by trypsin. These results represent an example of redox regulation of an enzymatic activity, which may play a role as a sensor of redox cellular status.

Foodborne Pathogens and Disease, 2019

Plant and essential oil extracts have been used for some time as antimicrobials and antioxidants,... more Plant and essential oil extracts have been used for some time as antimicrobials and antioxidants, although little is known about the interactions between the main components of these plant materials. This knowledge could help to design more potent antimicrobial and antioxidant mixtures. Carvacrol and thymol, the main components of the essential oils of the Lamiaceae family of plants, were assessed in combination to evaluate their antioxidant activity and antimicrobial effect against 19 strains of Staphylococcus aureus (S. aureus) of different origins (clinical, meat, milk, and other) and mostly (12) enterotoxin producers. The microdilution test assay was used to determine the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of the two phenolics alone and in combination. Based on the fractional inhibitory concentration index (FICI), no antimicrobial interaction (0.5 < FICI <4) between carvacrol and thymol was observed against 42% of the S. aureus strains and an antagonistic interaction (FICI >4) was observed in the rest, which indicates different behavior among strains in relation to this antimicrobial combination. Particularly, an antagonistic effect was observed in 29% of the meat origin strains and 57% of the dairy origin strains. Combinations of carvacrol and thymol were bactericidal (differences in MIC and MBC values not more than twofold) for 60% of the tested strains. At low concentrations of both components, the antioxidant effect is additive. However, at high concentrations (2.50 or 2.66 mM) of at least one of the components of the combination, it is antagonistic. The different types of interactions of the components in the combination can depend on many factors (ratio, structural characteristics, and the establishment of intermolecular complexes). The results could be used as reference to apply this combination in foods to control S. aureus, to maintain the organoleptic properties and to extend the shelf-life of them.

Biochemical Journal, 1982

1. Lactate dehydrogenase from mycelium of Phycomyces blakesleeanus showed positive homotropic int... more 1. Lactate dehydrogenase from mycelium of Phycomyces blakesleeanus showed positive homotropic interactions with NADH at all pH values studied (pH 5.0-7.7). The calculated values for the first and last intrinsic association constants remained unaltered with pH, in contrast with the Hill coefficient value, which varied significantly, reaching its maximum values at pH 6.0 and 7.7. This suggests the hypothesis that pH regulates these homotropic effects by changes in the value of the intermediate intrinsic association constants. 2. From pH 7.2 to 7.7 lactate dehydrogenase exhibited, likewise, positive homotropic interactions with pyruvate. There were practically no changes in the first and last intrinsic association constants and in Hill coefficient values with pH. At pH values below 7.2 (pH 5.0-6.8) the enzyme showed high substrate inhibition, which was highly dependent on pH, NADH concentration and temperature. By way of substrate inhibition pH regulates, primarily, lactate dehydrogena...

Journal of Enzyme Inhibition and Medicinal Chemistry, 1990

Studies on ATP effects on the allosteric kinetics shown by pyruvate kinase from Phycomyces blakes... more Studies on ATP effects on the allosteric kinetics shown by pyruvate kinase from Phycomyces blakesleeanus NRRL 1555 (-) are reported. Phosphoenolpyruvate showed an allosteric ATP-dependent substrate inhibition. The results supported the existence of spatially distinct catalytic binding sites and the inhibitory binding sites for phosphoenolpyruvate, and ATP showed opposite heterotropic effects with respect to these two types of binding site. With respect to Mg2+ ions, ATP caused a negative heterotropic effect. The global inhibitory effect of ATP was in agreement with the predictions postulated by the two-state concerted-symmetry model of Monod, Wyman and Changeux.

Biochemical Journal, 1990

Isocitrate lyase was purified from Phycomyces blakesleeanus N.R.R.L. 1555(-). The native enzyme h... more Isocitrate lyase was purified from Phycomyces blakesleeanus N.R.R.L. 1555(-). The native enzyme has an Mr of 240,000. The enzyme appeared to be a tetramer with apparently identical subunits of Mr 62,000. The enzyme requires Mg2+ for activity, and the data suggest that the Mg2(+)-isocitrate complex is the true substrate and that Mg2+ ions act as a non-essential activator. The kinetic mechanism of the enzyme was investigated by using product and dead-end inhibitors of the cleavage and condensation reactions. The data indicated an ordered Uni Bi mechanism and the kinetic constants of the model were calculated. The spectrophotometric titration of thiol groups in Phycomyces isocitrate lyase with 5.5′-dithiobis-(2-nitrobenzoic acid) gave two free thiol groups per subunit of enzyme in the native state and three in the denatured state. The isocitrate lyase was completely inactivated by iodoacetate, with non-linear kinetics. The inactivation data suggest that the enzyme has two classes of mo...

Research in Microbiology, 2005

Two forms of acetyl-CoA synthetase (ACS1 and ACS2) have been detected in Phycomyces blakesleeanus... more Two forms of acetyl-CoA synthetase (ACS1 and ACS2) have been detected in Phycomyces blakesleeanus. ACS1, encoded by the gene facA, was induced by acetate and repressed by glucose at the transcriptional level. ACS2, not encoded by the gene facA, was detected as a response to carbon starvation both in the wild type and in an facA − mutant. Both enzymes were purified and characterized. They can use acetate and propionate as substrates. ACS2 is a much more stable enzyme than ACS1. After 60 min incubation at 55 • C, ACS2 retained 50% of its activity whereas ACS1 only retained 3%. The optimum temperature was 50 • C for ACS2 and 30 • C for ACS1.

Biochimica et Biophysica Acta, 1988

Glutathione reductase (NAD(P)H:oxidized-glutathione oxidoreductase, EC 1.6.4.2) from the mycelium... more Glutathione reductase (NAD(P)H:oxidized-glutathione oxidoreductase, EC 1.6.4.2) from the mycelium of Phycomyces blakesleeanus NRRL 1555(−) was purified 7600-fold to electrophoretic homogeneity. The enzyme gave an absorption spectrum that is characteristic for a flavoprotein with and A 280 /A 460 of 5.5. The native enzyme molecule is a dimer ( M r 100 000 ) composed of identical subunits ( M r 50 200 ) each containing a FAD-bound molecule. The Stokes radius and D 20, w , were 38.3 A and 55.6 μm2/s, respectively. The optimum pH values were 7.5 and 6.0 with NADPH and NADH, respectively, as electron donors. Apparent K m values of 20.15 and 130 μM were determined at pH 7.5 for NADPH and GSSG, respectively. Substrate inhibition by GSSG was evident at acidic pH values. The Arrhenius plot of the enzymatic activity was linear with a value of the apparent activation energy of 8.6 kcal · mol−1. Enzyme activity was affected by the ionic strength of the assay medium. Phycomyces glutathione reduc...