Production of β-glucosidase and diauxic usage of sugar mixtures byCandida molischiana

1996 ◽  
Vol 42 (5) ◽  
pp. 431-436 ◽  
Author(s):  
Shelby N. Freer ◽  
Christopher D. Skory
Keyword(s):  
Enzyme Activity ◽  
Key Words ◽  
Yeast Species ◽  
Glucose Repression ◽  
Carbon Sources ◽  
Substrate Preference ◽  
Glucosidase Activity ◽  
Key Words Glucose ◽  
Sugar Mixtures

The fermentation of cellobiose is a rare trait among yeasts. Of the 308 yeast species that utilize cellobiose aerobically, only 12 species ferment it, and only 2 species, Candida molischiana and Candida wickerhamii, also ferment cellodextrins. Candida molischiana produced β-glucosidase activity on all carbon sources tested, except glucose, mannose, and fructose. When these sugars were added to cultures growing on cellobiose, the synthesis of β-glucosidase ceased. However, the total amount of enzyme activity remained constant, indicating that the C. molischiana β-glucosidase is catabolite repressed and not catabolite inactivated. When grown in medium initially containing glucose plus xylose, cellobiose, maltose, mannitol, or glucitol, C. molischiana preferentially utilized glucose and produced little β-glucosidase activity until glucose was nearly depleted from the medium. When grown in medium containing cellobiose plus either fructose or mannose, the yeast preferentially utilized the monosaccharides and produced little β-glucosidase activity. Candida molischiana produced β-glucosidase and co-utilized cellobiose and xylose, maltose, or trehalose. Glucose and fructose, mannose, or trehalose were co-utilized; however, no β-glucosidase activity was detected. Thus, the order of substrate preference groups appeared to be (glucose, trehalose, fructose, mannose) > (cellobiose, maltose, xylose) > (mannitol, glucitol).Key words: glucose repression, trehalase, diauxic utilization, yeast.

Utilization of glucose and cellobiose by Candida molischiana

1995 ◽  
Vol 41 (2) ◽  
pp. 177-185 ◽  
Author(s):  
S. N. Freer
Keyword(s):  
Enzyme Activity ◽  
Glucose Concentration ◽  
Glucose Repression ◽  
Carbon Sources ◽  
Soluble Starch ◽  
Complex Medium ◽  
Initial Glucose Concentration ◽  
Carbohydrate Source ◽  
Glucosidase Activity ◽  
Initial Ph

Some of the factors that influence the biosynthesis of the Candida molischiana β-glucosidase were investigated. The yeast produced maximal enzyme activity when grown at 28 °C in a carbohydrate-containing complex medium (YM) in which the initial pH was adjusted to 6.0. The enzyme appeared to be produced constitutively, as activity was detected when either ethanol, glycerol, xylose, glucitol, mannitol, maltose, trehalose, cellobiose, cellodextrins, or soluble starch was used as the carbohydrate source. The presence of either glucose, mannose, or fructose (> 25 g/L) repressed β-glucosidase expression; however, C. molischiana did produce β-glucosidase when the initial glucose concentration was <25 g/L. When the yeast was grown in YM medium containing glucose plus cellobiose, diauxic utilization of the carbon sources was observed, and β-glucosidase activity was not detected until the glucose was depleted from the medium.Key words: β-glucosidase, glucose repression, fermentation, yeast.

scholarly journals Multicopy FZF1 (SUL1) Suppresses the Sulfite Sensitivity but Not the Glucose Derepression or Aberrant Cell Morphology of a grr1 Mutant of Saccharomyces cerevisiae

Genetics ◽  
1996 ◽  
Vol 144 (2) ◽  
pp. 511-521 ◽  
Author(s):  
Dorina Avram ◽  
Alan T Bakalinsky
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Transcription Factor ◽  
Glucose Metabolism ◽  
Cell Morphology ◽  
Glucose Repression ◽  
Carbon Sources ◽  
Single Copy ◽  
Aberrant Cell ◽  
Growth Defects ◽  
Putative Transcription Factor

Abstract An ssu2 mutation in Sacccharomyces cermisiae, previously shown to cause sulfite sensitivity, was found to be allelic to GRR1, a gene previously implicated in glucose repression. The suppressor rgt1, which suppresses the growth defects of grr1 strains on glucose, did not fully suppress the sensitivity on glucose or nonglucose carbon sources, indicating that it is not strictly linked to a defect in glucose metabolism. Because the Cln1 protein was previously shown to be elevated in grr1 mutants, the effect of CLN1 overexpression on sulfite sensitivity was investigated. Overexpression in GRR1 cells resulted in sulfite sensitivity, suggesting a connection between CLN1 and sulfite metabolism. Multicopy FZF1, a putative transcription factor, was found to suppress the sulfite sensitive phenotype of grr1 strains, but not the glucose derepression or aberrant cell morphology. Multicopy FZF1 was also found to suppress the sensitivity of a number of other unrelated sulfite-sensitive mutants, but not that of ssu1 or met20, implying that FZF1 may act through Ssulp and Met20p. Disruption of FZF1 resulted in sulfite sensitivity when the construct was introduced in single copy at the FZF1 locus in a GRR1 strain, providing evidence that FZF1 is involved in sulfite metabolism.

Cloning and genetic mapping of SNF1, a gene required for expression of glucose-repressible genes in Saccharomyces cerevisiae

1984 ◽  
Vol 4 (1) ◽  
pp. 49-53
Author(s):  
J L Celenza ◽  
M Carlson
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Genetic Mapping ◽  
Gene Product ◽  
Structural Gene ◽  
Glucose Repression ◽  
Carbon Sources ◽  
Recombinant Plasmids ◽  
The Right ◽  
Integrating Vector

A functional SNF1 gene product is required to derepress expression of many glucose-repressible genes in Saccharomyces cerevisiae. Strains carrying a snf1 mutation are unable to grow on sucrose, galactose, maltose, melibiose, or nonfermentable carbon sources; utilization of these carbon sources is regulated by glucose repression. The inability of snf1 mutants to utilize sucrose results from failure to derepress expression of the structural gene for invertase at the RNA level. We isolated recombinant plasmids carrying the SNF1 gene by complementation of the snf1 defect in S. cerevisiae. A 3.5-kilobase region is common to the DNA segments cloned in five different plasmids. Transformation of S. cerevisiae with an integrating vector carrying a segment of the cloned DNA resulted in integration of the plasmid at the SNF1 locus. This result indicates that the cloned DNA is homologous to sequences at the SNF1 locus. By mapping a plasmid marker linked to SNF1 in this transformant, we showed that the SNF1 gene is located on chromosome IV. We then mapped snf1 to a position 5.6 centimorgans distal to rna3 on the right arm; snf1 is not extremely closely linked to any previously mapped mutation.

Regulation of yeast COX6 by the general transcription factor ABF1 and separate HAP2- and heme-responsive elements

1992 ◽  
Vol 12 (5) ◽  
pp. 2302-2314
Author(s):  
J D Trawick ◽  
N Kraut ◽  
F R Simon ◽  
R O Poyton
Keyword(s):  
Transcription Factor ◽  
Carbon Source ◽  
Protein Binding ◽  
Glucose Repression ◽  
Protein Complexes ◽  
Carbon Sources ◽  
Major Protein ◽  
Mobility Shift ◽  
Gel Mobility Shift ◽  
In Cells

Transcription of the Saccharomyces cerevisiae COX6 gene is regulated by heme and carbon source. It is also affected by the HAP2/3/4 transcription factor complex and by SNF1 and SSN6. Previously, we have shown that most of this regulation is mediated through UAS6, an 84-bp upstream activation segment of the COX6 promoter. In this study, by using linker scanning mutagenesis and protein binding assays, we have identified three elements within UAS6 and one element downstream of it that are important. Two of these, HDS1 (heme-dependent site 1; between -269 and -251 bp) and HDS2 (between -228 and -220 bp), mediate regulation of COX6 by heme. Both act negatively. The other two elements, domain 2 (between -279 and -269 bp) and domain 1 (between -302 and -281 bp), act positively. Domain 2 is required for optimal transcription in cells grown in repressing but not derepressing carbon sources. Domain 1 is essential for transcription per se in cells grown on repressing carbon sources, is required for optimal transcription in cells grown on a derepressing carbon source, is sufficient for glucose repression-derepression, and is the element of UAS6 at which HAP2 affects COX6 transcription. This element contains the major protein binding sites within UAS6. It has consensus binding sequences for ABF1 and HAP2. Gel mobility shift experiments show that domain 1 binds ABF1 and forms different numbers of DNA-protein complexes in extracts from cells grown in repressing or derepressing carbon sources. In contrast, gel mobility shift experiments have failed to reveal that HAP2 or HAP3 binds to domain 1 or that hap3 mutations affect the complexes bound to it. Together, these findings permit the following conclusions: COX6 transcription is regulated both positively and negatively; heme and carbon source exert their effects through different sites; domain 1 is absolutely essential for transcription on repressing carbon sources; ABF1 is a major component in the regulation of COX6 transcription; and the HAP2/3/4 complex most likely affects COX6 transcription indirectly.

Neutral α-glucosidase activity in mouse: a marker of epididymal function?

2007 ◽  
Vol 19 (4) ◽  
pp. 563 ◽  
Author(s):  
Ana C. Martini ◽  
Rosa I. Molina ◽  
Laura M. Vincenti ◽  
María E. Santillán ◽  
Graciela Stutz ◽  
...  
Keyword(s):  
Enzyme Activity ◽  
Food Restriction ◽  
Sperm Concentration ◽  
Secretory Activity ◽  
Spectrophotometric Analysis ◽  
Glucosidase Activity ◽  
Epididymal Maturation ◽  
Mouse Epididymis ◽  
Positive Controls

Neutral α-glucosidase (NAG) activity is considered a functional epididymal marker in several species. Unlike the rat, no NAG activity has been detected in mice. The aims of the present study were to evaluate NAG secretory activity (the supernatant of the incubated tissue) in mouse epididymis and to determine whether it could be used as a functional epididymal marker. Epididymides (whole or in parts) were incubated in the presence or absence of testosterone (10−5 m) and secretory NAG activity was compared with known positive controls. Furthermore, we compared enzyme activity in epididymides from well-fed and undernourished mice (50% food restriction for 21 days), a model that alters the epididymal maturation processes. Spectrophotometric analysis revealed NAG activity in mouse epididymis (22.6 ± 3.7 mU g–1 tissue; n = 4), being higher in the caput. NAG activity was statistically higher in the caput than in the corpus and in the cauda. No significant differences existed between the caput NAG activity and complete epididymis NAG activity. In undernourished mice, we confirmed changes in epididymal maturation observed previously (i.e. increased number of immature spermatozoa and diminution of the sperm concentration). Concordantly, the epididymides of undernourished mice exhibited decreased enzyme secretory activity, which increased to values similar to those seen in controls following incubation in the presence of testosterone (22.5 ± 2.6, 12.5 ± 1.0 and 22.4 ± 3.7 mU g–1 tissue, n = 9 in control (n = 7), undernourished (n = 9) and undernourished + testosterone groups (n = 9), respectively). In conclusion, NAG activity was detected in mouse epididymis. Although the present study supports the possibility of using NAG as an epididymal marker, more studies are necessary to effectively prove that NAG activity can be used as an epididymal marker.

scholarly journals Re-evaluation of Mature Seed-derived Callusing and Regeneration Potential of Nine Orchardgrass (Dactylis golomerata L.) Cultivars

1970 ◽  
Vol 17 (2) ◽  
pp. 193-207 ◽  
Author(s):  
Sang-Hoon Lee ◽  
Nagib Ahsan ◽  
Ki-Won Lee ◽  
Dong-Gi Lee ◽  
Iftekhar Alam ◽  
...  
Keyword(s):  
Key Words ◽  
Callus Induction ◽  
Plant Tissue ◽  
Genotypic Variation ◽  
Carbon Sources ◽  
Mature Seed ◽  
Regeneration Potential ◽  
Efficient Regeneration ◽  
N6 Medium ◽  
Mature Seeds

A suitable callus induction and efficient regeneration protocol for orchardgrass (Dactylis golomerata L.) was developed. It consisted of 3 mg/l 2,4-D + 0.1 mg/l BA + 1 g/l CH + 300 mg/l L-proline + 40 mg/l L-cysteine + 30 g/l sucrose in MS showed the highest percentage of callus induction. Maltose exhibited better in regeneration than other types of carbon sources. Highest (71%) regeneration was obtained from N6 medium containing 1 mg/l 2,4-D + 3 mg/l BA + 1 g/l CH + 300 mg/l L-proline + 40 mg/l L-cysteine + 30 g/l maltose. Among the nine cultivars of orchardgrass (Dactylis golomerata L.), genotypic variation was observed in both callus induction and regeneration. Overall callus induction and regeneration rates were 23 - 73 and 17 - 71%, respectively.  Key words: Dactylis golomerata, Orchardgrass, Mature seeds, Additives, Regeneration, Maltose. D.O.I. 10.3329/ptcb.v17i2.3240 Plant Tissue Cult. & Biotech. 17(2): 193-207, 2007 (December)

scholarly journals Divergent MLS1 promoters lie on a fitness plateau for gene expression

2015 ◽  
Author(s):  
Andrew C Bergen ◽  
Gerilyn M Olsen ◽  
Justin C Fay
Keyword(s):  
Gene Expression ◽  
Saccharomyces Cerevisiae ◽  
Dynamic Response ◽  
Yeast Species ◽  
Glucose Repression ◽  
Gene Activation ◽  
Malate Synthase ◽  
Expression Divergence ◽  
Fitness Effects ◽  
Single Promoter

Qualitative patterns of gene activation and repression are often conserved despite an abundance of quantitative variation in expression levels within and between species. A major challenge to interpreting patterns of expression divergence is knowing which changes in gene expression affect fitness. To characterize the fitness effects of gene expression divergence we placed orthologous promoters from eight yeast species upstream of malate synthase (MLS1) in Saccharomyces cerevisiae. As expected, we found these promoters varied in their expression level under activated and repressed conditions as well as in their dynamic response following loss of glucose repression. Despite these differences, only a single promoter driving near basal levels of expression caused a detectable loss of fitness. We conclude that the MLS1 promoter lies on a fitness plateau whereby even large changes in gene expression can be tolerated without a substantial loss of fitness.

scholarly journals Regulation of yeast COX6 by the general transcription factor ABF1 and separate HAP2- and heme-responsive elements.

1992 ◽  
Vol 12 (5) ◽  
pp. 2302-2314 ◽  
Author(s):  
J D Trawick ◽  
N Kraut ◽  
F R Simon ◽  
R O Poyton
Keyword(s):  
Transcription Factor ◽  
Carbon Source ◽  
Protein Binding ◽  
Glucose Repression ◽  
Protein Complexes ◽  
Carbon Sources ◽  
Major Protein ◽  
Mobility Shift ◽  
Gel Mobility Shift ◽  
In Cells

Transcription of the Saccharomyces cerevisiae COX6 gene is regulated by heme and carbon source. It is also affected by the HAP2/3/4 transcription factor complex and by SNF1 and SSN6. Previously, we have shown that most of this regulation is mediated through UAS6, an 84-bp upstream activation segment of the COX6 promoter. In this study, by using linker scanning mutagenesis and protein binding assays, we have identified three elements within UAS6 and one element downstream of it that are important. Two of these, HDS1 (heme-dependent site 1; between -269 and -251 bp) and HDS2 (between -228 and -220 bp), mediate regulation of COX6 by heme. Both act negatively. The other two elements, domain 2 (between -279 and -269 bp) and domain 1 (between -302 and -281 bp), act positively. Domain 2 is required for optimal transcription in cells grown in repressing but not derepressing carbon sources. Domain 1 is essential for transcription per se in cells grown on repressing carbon sources, is required for optimal transcription in cells grown on a derepressing carbon source, is sufficient for glucose repression-derepression, and is the element of UAS6 at which HAP2 affects COX6 transcription. This element contains the major protein binding sites within UAS6. It has consensus binding sequences for ABF1 and HAP2. Gel mobility shift experiments show that domain 1 binds ABF1 and forms different numbers of DNA-protein complexes in extracts from cells grown in repressing or derepressing carbon sources. In contrast, gel mobility shift experiments have failed to reveal that HAP2 or HAP3 binds to domain 1 or that hap3 mutations affect the complexes bound to it. Together, these findings permit the following conclusions: COX6 transcription is regulated both positively and negatively; heme and carbon source exert their effects through different sites; domain 1 is absolutely essential for transcription on repressing carbon sources; ABF1 is a major component in the regulation of COX6 transcription; and the HAP2/3/4 complex most likely affects COX6 transcription indirectly.

scholarly journals Functional domains in the Mig1 repressor.

1996 ◽  
Vol 16 (3) ◽  
pp. 753-761 ◽  
Author(s):  
J Ostling ◽  
M Carlberg ◽  
H Ronne
Keyword(s):  
Zinc Finger Protein ◽  
Wilms Tumor ◽  
Glucose Repression ◽  
Protein A ◽  
Carbon Sources ◽  
Negative Control ◽  
Functional Domains ◽  
Deletion Mapping ◽  
Yeast Saccharomyces Cerevisiae ◽  
Finger Protein

Mig1 is a zinc finger protein that mediates glucose repression in the yeast Saccharomyces cerevisiae. It is related to the mammalian Krox/Egr, Wilms' tumor, and Sp1 proteins and binds to a GC-rich motif that resembles the GC boxes recognized by these proteins. We have performed deletion mapping in order to identify functional domains in Mig1. We found that a small C-terminal domain comprising the last 24 amino acids mediates Mig1-dependent repression of a reporter gene. This effector domain contains several leucine-proline dipeptide repeats. We further found that inhibition of Mig1 activity in the absence of glucose is mediated by two internal elements in the Mig1 protein. A Mig1-VP16 hybrid activator was used to further investigate how Mig1 is regulated. Mig1-VP16 can activate transcription from promoters containing Mig1-binding sites and suppresses the inability of Snf1-deficient cells to grow on certain carbon sources. We found that a deletion of the SNF1 gene increases the activity of Mig1-VP16 fivefold under derepressing conditions but not in the presence of glucose. This shows that the hybrid activator is under negative control by the Snf1 protein kinase. Deletion mapping within Mig1-VP16 revealed that regulation of its activity by Snf1 is conferred by the same internal elements in the Mig1 sequence that mediate inhibition of Mig1 activity in the absence of glucose.

scholarly journals Duplicate upstream activating sequences in the promoter region of the Saccharomyces cerevisiae GAL7 gene.

1986 ◽  
Vol 6 (1) ◽  
pp. 246-256 ◽  
Author(s):  
M Tajima ◽  
Y Nogi ◽  
T Fukasawa
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Glucose Repression ◽  
Rotational Symmetry ◽  
Consensus Sequence ◽  
Carbon Sources ◽  
Constitutive Expression ◽  
Tata Box ◽  
Base Pairs ◽  
Multicopy Vector ◽  
In Cis

We constructed a series of deletions in the 5' noncoding region of the Saccharomyces cerevisiae GAL7 gene, fused them to the Escherichia coli gene lacZ, and introduced them into yeasts by using a multicopy vector. We then studied the effect of the deletions on beta-galactosidase synthesis directed by the gene fusions in media with various carbon sources. This analysis identified a TATA box and two upstream activating sequences as necessary elements for galactose-controlled GAL7 transcription. Two upstream activating sequences exhibiting 71% homology with each other were located 255 and 168 base pairs, respectively, upstream of the GAL7 transcription start point. Each sequence consists of 21 base pairs, displaying an approximate rotational symmetry with a core consensus sequence of GAA--AGCTGCTTC--CGCG. At least one of the two sequences is required for galactose induction and also for glucose repression of the GAL7'-lac'Z gene. Analysis with host regulatory mutants delta gal14 and delta gal180 suggests that these sequences are the site at which the GAL4 product exerts its action to activate the GAL7 gene. We also observed that a deletion lacking both upstream activation sequences allowed the gene fusion to be expressed in the absence of galactose at about 10% of the fully induced level of the intact fusion. This constitutive expression depended on the presence of the TATA box of GAL7 in cis but not on a functional GAL4 gene. The level of the uncontrolled expression was decreased by increasing the distance between the TATA box and the pBR322 sequence in the vector plasmid.

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