Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Ethanol utilization”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 109 records · Page 6Linked to original sources
PubMed1995

The basal level of transcription of the alc genes in the ethanol regulon in Aspergillus nidulans is controlled both by the specific transactivator AlcR and the general carbon catabolite repressor CreA.

In the A. nidulans ethanol utilization pathway, specific induction is mediated by the transactivator AlcR which is subject to strong positive autogenous regulation and activates the transcription of the two structural genes alcA and aldA. Carbon catabolite repression is mediated by CreA which represses directly the transacting gene alcR and the two structural genes. We show here that the basal expression of the alcR and alcA genes is also controlled by the two regulatory circuits, positively by the transactivator AlcR and negatively by the repressor CreA, the aldA gene being subject only to the control of the CreA repressor.

Aspergillus nidulans↗
PubMed1995

IMP2, a gene involved in the expression of glucose-repressible genes in Saccharomyces cerevisiae.

Two mutants carrying different deletions of the IMP2 coding sequence of Saccharomyces cerevisiae, delta T1, which encodes a protein lacking the last 26 C-terminal amino acids, and delta T2, which completely lacks the coding region, were analysed for derepression of glucose-repressible maltose, galactose, raffinose and ethanol utilization pathways in response to glucose limitation. The role of the IMP2 gene product in the regulation of carbon catabolite repressible enzymes maltase, invertase, alcohol dehydrogenase, NAD-dependent glutamate dehydrogenase (NAD-GDH) and L-lactate:ferricytochrome-c oxidoreductase (L-LCR) was also analysed. The IMP2 gene product is required for the rapid glucose derepression of all above-mentioned carbon source utilization pathways and of all the enzymes except for L-LCR. NAD-GDH is regulated by IMP2 in the opposite way and, in fact, this enzyme was released at higher levels in both imp2 mutants than in the wild-type strain. Therefore, the product of IMP2 appears to be involved in positive and negative regulation. Both deletions result in growth and catalytic defects; in some cases partial modification of the gene product yielded more dramatic effects than its complete absence. Moreover, evidence is provided that the IMP2 gene product regulates galactose- and maltose-inducible genes at the transcriptional level and is a positive regulator of maltase, maltose permease and galactose permease gene expression.

Alleles↗
PubMed1996

A newly identified gene cluster in Aspergillus nidulans comprises five novel genes localized in the alc region that are controlled both by the specific transactivator AlcR and the general carbon-catabolite repressor CreA.

Ethanol-utilization in Aspergillus nidulans is mediated by alcohol dehydrogenase I and aldehyde dehydrogenase encoded by alcA and aldA, respectively. Both genes are under the transcriptional control of the specific activator AlcR and the general carbon catabolite repressor CreA. The alcR and alcA genes are closely linked in chromosome VII; aldA is located in chromosome VIII. We have identified five other transcripts that are expressed from the same genomic region as alcA and alcR. They are inducible by the gratuitous inducer ethyl methyl ketone (EMK), and are carbon catabolite repressed. The corresponding genes, designated alcM, alcS, alcO, alcP, and alcU, are differentially regulated by the specific transcriptional activator AlcR, and they are not all under the direct control by the CreA repressor. Some of the inducible transcripts are very abundant in the cell, whereas others are poorly expressed. Two sets of genes, alcM/alcS and alcR/alcO, are divergently transcribed and probably share a common cis-acting region, whereas alcP and alcU are individually transcribed from the same strand as alcA and alcR, and have their own promoters. The significance of the alc gene clustering is discussed. At least four of the five novel alc genes in the cluster are not essential for ethanol metabolism.

Aspergillus nidulans↗
PubMed1994

The capacity of the malate-aspartate shuttle differs between periportal and perivenous hepatocytes from rats.

The capacity of the malate-aspartate shuttle was evaluated in periportal (PP-H) and perivenous subfraction of rat hepatocytes (PV-H). The rate of glutamine production from alanine was 34-fold higher in PV-H than in PP-H. Statistically significant differences between PP-H and PV-H were found for the activities of lactate dehydrogenase and pyruvate kinase but not for the activities of NAD(+)-malate dehydrogenase, aspartate aminotransferase, and mitochondrial alanine aminotransferase. The rate of glucose production from sorbitol and the rate of ethanol utilization were higher in PP-H than in PV-H. In the presence of phenazine methosulfate (PMS), the increments in these rates were significantly greater in PV-H than in PP-H. The capacity of malate-aspartate shuttle in the presence of alanine was significantly higher in PP-H than in PV-H but in the presence of asparagine was similar in PP-H and PV-H. The results suggest that the capacity of malate-aspartate shuttle distributes heterogeneously along liver lobules with the dominance in periportal zone and that the difference of the capacity may result from the difference in the transport of aspartate across the mitochondrial membrane.

Alanine↗
PubMed1992

Identification of the promoter region involved in the autoregulation of the transcriptional activator ALCR in Aspergillus nidulans.

The ALCR protein is the transcriptional activator of the ethanol utilization pathway in the filamentous fungus Aspergillus nidulans. This activator belongs to a family of fungal proteins having a conserved DNA-binding domain containing six cysteines (C6 class) with some striking features. At variance with other motifs of this class, the binding domain of ALCR is strongly asymmetrical in relation to the central cysteines and moreover was predicted to adopt a helix-turn-helix structure. This domain of ALCR was synthesized in Escherichia coli and purified as a glutathione-S-transferase fusion protein. Our results show that the transcriptional activator ALCR is a DNA-binding protein. The DNA-binding motif contains zinc that is necessary for the specific DNA binding. The ALCR peptide binds upstream of the coding region of alcR to two specific targets with different affinities that are characterized by a conserved 5-nucleotide core, 5'-CCGCA-3' (or its reverse). One site, the lower-affinity binding site, is a direct repeat, and the other, the higher-affinity binding site, is a palindromic sequence with dyad symmetry. Therefore, the ALCR binding protein is able to recognize one DNA sequence in two different configurations. An alcR mutant obtained by deletion of the two specific targets in the cis-acting region of the alcR gene is unable to grow on ethanol and does not express any alcohol dehydrogenase activity. These results demonstrate that the binding sites are in vivo functional targets (UASalc) for the ALCR protein in A. nidulans. They corroborate prior evidence that alcR is autoregulated.

Aspergillus nidulans↗
PubMed1998

The CreA repressor is the sole DNA-binding protein responsible for carbon catabolite repression of the alcA gene in Aspergillus nidulans via its binding to a couple of specific sites.

Carbon catabolite repression is mediated in Aspergillus nidulans by the negative acting protein CreA. The CreA repressor plays a major role in the control of the expression of the alc regulon, encoding proteins required for the ethanol utilization pathway. It represses directly, at the transcriptional level, the specific transacting gene alcR, the two structural genes alcA and aldA, and other alc genes in all physiological growth conditions. Among the seven putative CreA sites identified in the alcA promoter region, we have determined the CreA functional targets in AlcR constitutive and derepressed genetic backgrounds. Two different divergent CreA sites, of which one overlaps a functional AlcR inverted repeat site, are largely responsible for alcA repression. Totally derepressed alcA expression is achieved when these two CreA sites are disrupted in addition to another single site, which overlaps the functional palindromic induction target. The fact that derepression is always associated with alcA overexpression is consistent with a competition model between AlcR and CreA for their cognate targets in the same region of the alcA promoter. Our results also indicate that the CreA repressor is necessary and sufficient for the total repression of the alcA gene.

Alcohol Dehydrogenase↗
PubMed1992

Specific binding sites for the activator protein, ALCR, in the alcA promoter of the ethanol regulon of Aspergillus nidulans.

ALCR is the specific activator of the Aspergillus nidulans ethanol-utilization pathway, mediating the induction of its own transcription and that of the structural genes alcA and aldA, encoding respectively, alcohol dehydrogenase I and aldehyde dehydrogenase. ALCR is a DNA binding protein in which 6 cysteines are coordinated in a zinc binuclear cluster. This domain was fused to glutathione-S-transferase (GST) and isolated as a GST-ALCR(7-58*) fusion protein from Escherichia coli. Mobility shift assays showed that the ALCR fusion protein binds at sites upstream of the alcA promoter. DNaseI protection footprinting experiments revealed three specific binding sites, two that are direct repeats and one that is an inverted repeat with the same half-site 5'-CCGCA-3'. The half-sites are separated by a variable number of nucleotides in both types of target. The interaction of the ALCR fusion protein with direct and inverted repeats were examined by using interference and protection footprinting assays. In both binding sites, modification of the guanines in the half-sites interfered with the formation of the DNA complex, but the adjacent ones did not. Our results suggest that the ALCR protein makes contact in the major groove of the DNA helix of the half-sites. The functionality of two out of three binding sites of the GST-ALCR protein was demonstrated after their deletion. Therefore, the region encompassing these binding sites is a cis-acting element involved in the full induction of the alcA gene.

Alcohol Dehydrogenase↗
PubMed2000

In vivo studies of upstream regulatory cis-acting elements of the alcR gene encoding the transactivator of the ethanol regulon in Aspergillus nidulans.

The alcR gene of Aspergillus nidulans, which encodes the specific transactivator of the ethanol utilization pathway, is positively autoregulated and carbon catabolite repressed. Regulation by these two circuits occurs at the transcriptional level via the binding of the two regulators, AlcR and CreA, to their cognate targets respectively. We demonstrate here that out of two clustered putative AlcR repeated consensus sequences, only the palindromic target is functional in vivo. Hence, it is solely responsible for the alcR positive autogenous activation loop. Transcript mapping of the alcR gene showed that transcription initiation can occur at 553 bp and at or near 86 bp upstream of the start codon. These transcription start sites yield a transcript of 3.0 kb, which appears only under induced growth conditions, and of 2.6 kb, which is present under both induced and non-induced growth conditions respectively. Nine CreA consensus sites are present in the alcR promoter but only two pairs of two sites are functional in vivo. One of them is located in close proximity to the AlcR functional target. Within this pair, both sites are necessary to mediate a partial repression of alcR transcription. Disruption of either site results in an overexpression of alcR due to the absence of direct competition between AlcR and CreA for the same DNA region. The second functional pair of CreA sites is located between the two transcription initiation sites. Disruption of either of the two sites results in a totally derepressed alcR transcription, showing that they work as a pair constituting the more efficient repression mechanism. Thus, CreA acts by two different mechanisms: by competing with AlcR for the same DNA region and by an efficient direct repression. The latter mechanism presumably interfers with the general transcriptional machinery.

Aspergillus nidulans↗
PubMed2000

A HPLC-fluorescence detection method for determination of cardiac phospholipase D activity in vitro.

A nonradioactive assay for the investigation of phospholipase D (PLD) activity in cardiac membranes has been developed. A fluorescent derivative of phosphatidylcholine [2-decanoyl-1-(O-(11-(4,4-difluoro-5,7-dimethyl-4-bora-3a, 4a-diaza-s-indacene-3proprionyl)amino) undecyl) sn-glycero-3-phosphocholine] was utilized as substrate in an in vitro PLD-catalyzed transphosphatidylation reaction utilizing ethanol as second substrate. Unreacted phosphatidylcholine and the products of phospholipase activity (PEtOH, phosphatidylethanol; PA, phosphatidic acid; DAG, diacylglycerol) were separated by a binary gradient HPLC system and detected by fluorometry. The detection limit of this assay is approximately 0.6 pmol PEtOH. The reaction proceeded at a linear rate for up to 45 min and increased linearly with increasing amounts of rat cardiac membrane protein in a range of 0.625 microg up to at least 25 microg. In the presence of potassium fluoride, formation of fluorescent PA increased at the expense of DAG generation, demonstrating the presence of PA phosphohydrolase activity in rat cardiac membranes. PEtOH formation was unchanged in the presence of the PA phosphohydrolase inhibitor, indicating that the phosphatidylalcohol is not subject to further metabolism by this enzyme. Activation of protein kinase C by phorbol ester significantly increased PLD activity in cardiac membranes. This assay proved to be sensitive for accurate and rapid assessment of PLD activity in cardiac membranes permitting further characterization of the regulation of PLD signal transduction in the heart.

Animals↗
PubMed2001

ADHII in Aspergillus nidulans is induced by carbon starvation stress.

In Aspergillus nidulans there are three NAD(+)-dependent alcohol dehydrogenases (ADHs) that are capable of utilizing ethanol as a substrate. ADHI is the physiological enzyme of ethanol catabolism and ADHIII is induced under conditions of anaerobiosis. The physiological role of ADHII (structural gene alcB) is unknown. We have measured beta-galactosidase in a transformant with an alcB::lacZ fusion and have shown that alcB is maximally expressed under conditions of carbon starvation. The behavior of the alcB::lacZ transformant suggests a hierarchy of repressing carbon sources characteristic of repression by the general carbon catabolite repressor protein, CreA, but in a creA(d)30 background the transformant shows only partial derepression of beta-galactosidase on 1% glucose compared to the creA+ strain. Our results suggest that, in addition to carbon catabolite repression acting via CreA, a CreA-independent mechanism is involved in induction of alcB on carbon starvation.

Alcohol Dehydrogenase↗
PubMed2006

The natural history of Drosophila ehrmanae.

Drosophila ehrmanae [Parsons P. A., and Bock I. R. (1977). Aust. J. Zool. 25: 249-268] is a desiccation-sensitive species of undisturbed southern Australian temperate-zone rainforests. In this habitat category, nine Drosophila species have been collected, all belonging to the dominant Australian subgenus, Scaptodrosophila. Adults of these species are collected by sweeping foliage in damp habitats so providing temperature/desiccation information on adaptive behavioral selection within microhabitats. Biochemical and metabolic analyses of two common species of this subgenus indicate that D. ehrmanae should be alcohol dehydrogenase null with low ADH activity, should not utilize ethanol and its derivatives as a major resource, and therefore should not be attracted to these metabolites. Ultimately, such species may offer possibilities for behavioral/genetic analyses.

Animals↗
PubMed1987

Pyruvate carboxylase in the yeast pyc mutant.

Pyruvate carboxylase deficiency was previously reported to be the biochemical lesion in a yeast mutant, designated pyc, which cannot utilize ethanol, acetate, pyruvate, aspartate, or oxaloacetate as the sole carbon source [C. Wills and T. Melham (1985) Arch. Biochem. Biophys. 236, 782-791; C. Wills et al. (1986) Arch. Biochem. Biophys. 246, 306-320]. We present evidence here that the level of pyruvate carboxylase activity as well as the native and subunit molecular weights of this enzyme are identical in the mutant and the wild type. In addition we have used immunocytochemical labeling to demonstrate the exclusively cytosolic localization of this enzyme in both the mutant and wild-type yeast.

Carbon Dioxide↗
PubMed1995

Structure of simusan, a new acidic exopolysaccharide from Arthrobacter sp.

Simusan, a major exopolysaccharide produced by an ethanol-utilizing Arthrobacter sp. strain CE-17, contains D-glucose, D-mannose, D-galactose, L-rhamnose, D-glucuronic acid, and pyruvic acid in the ratios approximately 3:2:1:1:1:1 as well as O-acyl groups (presumably (presumably residues of acetic and palmitic acid). On the basis of chemical modifications of the polysaccharide, solvolysis with anhydrous hydrogen fluoride resulting in a penta- and an octa-saccharide fragment, Smith degradation, and 1H and 13C NMR analysis, the following structure of the repeating unit was established: [formula: see text] It is suggested that at least one of the glucose residues and the galactose residue are O-acetylated.

Arthrobacter↗
PubMed1994

Chromate-mediated free radical generation from cysteine, penicillamine, hydrogen peroxide, and lipid hydroperoxides.

The Cr(VI)-mediated free radical generation from cysteine, penicillamine, hydrogen peroxide, and model lipid hydroperoxides was investigated utilizing the electron spin resonance (ESR) spin trapping technique. Incubation of Cr(VI) with cysteine (Cys) generated cysteinyl radical. Radical yield depended on the relative concentrations of Cr(VI) and Cys. The radical generation became detectable at a cysteine:Cr(VI) ratio of about 5, reached its highest level at a ratio of 30, and declined thereafter. Cr(VI) or Cys alone did not generate a detectable amount of free radicals. Similar results were obtained with penicillamine. Incubation of Cr(VI), Cys or penicillamine and H2O2 led to hydroxyl (.OH) radical generation, which was verified by quantitative competition experiments utilizing ethanol. The mechanism for .OH radical generation is considered to be a Cr(VI)-mediated Fenton-like reaction. When model lipid hydroperoxides such as t-butyl hydroperoxide and cumene hydroperoxide were used in place of H2O2, hydroperoxide-derived free radicals were produced. Since thiols, such as Cys, exist in cellular systems at relatively high concentrations, Cr(VI)-mediated free radical generation in the presence of thiols may participate in the mechanisms of Cr(VI)-induced toxicity and carcinogenesis.

Chromates↗
PubMed1997

First experimental evidence of a zinc binuclear cluster in AlcR protein, mutational and X-ray absorption studies.

AlcR is the transcriptional activator of the ethanol utilization pathway in Aspergillus nidulans. The zinc DNA-binding domain contains ligands of zinc, six cysteines (Zn2Cys6) or five cysteines and one histidine (Zn2Cys5His). The utilisation of complementary approaches such as X-ray absorption spectroscopy, mutational analysis, zinc content evaluation, determination of specific binding connecting structural and biological data, have allowed to determine zinc environment and to analyse the involvement of amino acids. The determination by EXAFS of zinc ligands (four sulphur atoms), the Zn content in the protein (2:1), the evaluation of the distance between two zinc atoms (3.16 +/- 0.02 angstroms), together with the total loss of specific DNA-binding activity when one cysteine ligand is mutated, are in favour of a zinc cluster model in which six cysteine sulphurs ligate two zinc atoms. XANES spectra of wild type and H10A AlcR protein are virtually identical indicating that Histidine 10 does not have a direct contribution in zinc ligation but electrophoretic mobility shift assays show that His10 is involved in DNA-binding. In contrast, proline 25 does not seem to play any direct role in the DNA-binding activity but XANES spectra of Pro25A AlcR protein are slightly modified comparing to the wild type protein spectra. This suggests a role of the proline in the stabilisation of the Zn cluster structure. AlcR DNA-binding domain belongs to the zinc binuclear class family (Zn2Cys6) with unique characteristics resulting from its primary and secondary structures and its binding specificity toward direct and inverted repeat target.

Amino Acid Sequence↗
PubMedDate not supplied

CO2 and fluorinated solvent-based technologies for protein microparticle precipitation from aqueous solutions.

Precipitation with a compressed or supercritical fluid antisolvent (PCA) has been used to produce microparticles of biologically active proteins, pharmaceuticals, and polymers. However, the application of PCA to a wider range of proteins is limited by the low mutual solubility of water (necessary to dissolve most proteins) and CO(2) (traditionally used as the compressed antisolvent). This investigation extends PCA to proteins in aqueous solutions by utilizing ethanol as a cosolvent to enhance the antisolvent properties of CO(2) toward aqueous systems. alpha-Chymotrypsin, a model protein, was precipitated from both compressed CO(2) and a liquid fluorinated antisolvent, a hydrofluoroether (HFE). The equilibrium phase behavior of the antisolvent/ethanol/water systems was examined to identify a one-phase region suitable for protein precipitation. Spherical protein microparticles with a primary particle size of approximately 0.2-0.6 microm were recovered using both the compressed CO(2) and fluorinated antisolvents. Although the proteins retained significant activity using both antisolvent systems, the HFE-precipitated chymotrypsin retained higher activity than the CO(2)-precipitated protein.

Carbon Dioxide↗
PubMed1997

Maize glutathione-dependent formaldehyde dehydrogenase cDNA: a novel plant gene of detoxification.

We have previously shown that intact plants and cultured plant cells can metabolize and detoxify formaldehyde through the action of a glutathione-dependent formaldehyde dehydrogenase (FDH), followed by C-1 metabolism of the initial metabolite (formic acid). The cloning and heterologous expression of a cDNA for the glutathione-dependent formaldehyde dehydrogenase from Zea mays L. is now described. The functional expression of the maize cDNA in Escherichia coli proved that the cloned enzyme catalyses the NAD(+)- and glutathione (GSH)-dependent oxidation of formaldehyde. The deduced amino acid sequence of 41 kDa was on average 65% identical with class III alcohol dehydrogenase from animals and less than 60% identical with conventional plant alcohol dehydrogenases (ADH) utilizing ethanol. Genomic analysis suggested the existence of a single gene for this cDNA. Phylogenetic analysis supports the convergent evolution of ethanol-consuming ADHs in animals and plants from formaldehyde-detoxifying ancestors. The high structural conservation of present-day glutathione-dependent FDH in microorganisms, plants and animals is consistent with a universal importance of these detoxifying enzymes.

Alcohol Dehydrogenase↗
PubMed2001

RcoA has pleiotropic effects on Aspergillus nidulans cellular development.

Aspergillus nidulans rcoA encodes a member of the WD repeat family of proteins. The RcoA protein shares sequence similarity with other members of this protein family, including the Saccharomyces cerevisiae Tup1p and Neurospora crassa RCO1. Tup1p is involved in negative regulation of an array of functions including carbon catabolite repression. RCO1 functions in regulating pleiotropic developmental processes, but not carbon catabolite repression. In A. nidulans, deletion of rcoA (DeltarcoA), a recessive mutation, resulted in gross defects in vegetative growth, asexual spore production and sterigmatocystin (ST) biosynthesis. Expression of the asexual and ST pathway-specific regulatory genes, brlA and aflR, respectively, but not the signal transduction genes (i.e. flbA, fluG or fadA) regulating brlA and aflR expression was delayed (brlA) or eliminated (aflR) in a DeltarcoA strain. Overexpression of aflR in a DeltarcoA strain could not rescue normal expression of downstream targets of AflR. CreA-dependent carbon catabolite repression of starch and ethanol utilization was only weakly affected in a DeltarcoA strain. The strong role of RcoA in development, vegetative growth and ST production, compared with a relatively weak role in carbon catabolite repression, is similar to the role of RCO1 in N. crassa.

Amino Acid Sequence↗