Induction and repression of L-fucose dehydrogenase of Pullularia pullulans

The growth of Pullularia pullulans on L-fucose as the sole carbon source induces the synthesis of L-fucose dehydrogenase, a NAD-dependent enzyme that catalyzes the oxidation of L-fucose to L-fucono-δ-lactone, which spontaneously hydrolyzes to L-fuconic acid. The induction of the enzyme is inhibited by cycloheximide, suggesting de novo synthesis. D-Glucose, D-galactose, and glycerol at 0.5% concentration gave rise to 62, 54, 51, and 46% of repression of enzyme synthesis, respectively. No repression effect was detected with D-arabinose and L-rhamnose. L-Arabinose repressed only 20% of the enzyme synthesis. Among the sugars tested, only L-fucose and L-galactose were oxidized by the enzyme. L-Galactose, which shares a common ring structure with L-fucose, showed only 10% of the activity observed when L-fucose was used.

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N-Acetylglucosamine Utilization by Saccharomyces cerevisiae Based on Expression of Candida albicans NAG Genes

Applied and Environmental Microbiology ◽

10.1128/aem.00053-09 ◽

2009 ◽

Vol 75 (18) ◽

pp. 5840-5845 ◽

Cited By ~ 34

Author(s):

Jürgen Wendland ◽

Yvonne Schaub ◽

Andrea Walther

Keyword(s):

Saccharomyces Cerevisiae ◽

Candida Albicans ◽

Carbon Source ◽

Sole Carbon Source ◽

De Novo ◽

Biofuel Production ◽

Bioethanol Production ◽

Catabolic Pathway ◽

Cellular Functions ◽

Kinase A

ABSTRACT Synthesis of chitin de novo from glucose involves a linear pathway in Saccharomyces cerevisiae. Several of the pathway genes, including GNA1, are essential. Genes for chitin catabolism are absent in S. cerevisiae. Therefore, S. cerevisiae cannot use chitin as a carbon source. Chitin is the second most abundant polysaccharide after cellulose and consists of N-acetylglucosamine (GlcNAc) moieties. Here, we have generated S. cerevisiae strains that are able to use GlcNAc as a carbon source by expressing four Candida albicans genes (NAG3 or its NAG4 paralog, NAG5, NAG2, and NAG1) encoding a GlcNAc permease, a GlcNAc kinase, a GlcNAc-6-phosphate deacetylase, and a glucosamine-6-phosphate deaminase, respectively. Expression of NAG3 and NAG5 or NAG4 and NAG5 in S. cerevisiae resulted in strains in which the otherwise-essential ScGNA1 could be deleted. These strains required the presence of GlcNAc in the medium, indicating that uptake of GlcNAc and its phosphorylation were achieved. Expression of all four NAG genes produced strains that could use GlcNAc as the sole carbon source for growth. Utilization of a GlcNAc catabolic pathway for bioethanol production using these strains was tested. However, fermentation was slow and yielded only minor amounts of ethanol (approximately 3.0 g/liter), suggesting that fructose-6-phosphate produced from GlcNAc under these conditions is largely consumed to maintain cellular functions and promote growth. Our results present the first step toward tapping a novel, renewable carbon source for biofuel production.

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Regulation of pyrimidine formation in Pseudomonas lundensis

Canadian Journal of Microbiology ◽

10.1139/w08-137 ◽

2009 ◽

Vol 55 (3) ◽

pp. 261-268 ◽

Cited By ~ 1

Author(s):

Thomas P. West

Keyword(s):

Carbon Source ◽

Orotic Acid ◽

De Novo ◽

Enzyme Synthesis ◽

Chemical Mutagenesis ◽

Aspartate Transcarbamoylase ◽

Food Spoilage ◽

Orotate Phosphoribosyltransferase ◽

Selection Procedures ◽

Monophosphate Decarboxylase

The regulation of pyrimidine formation in the food spoilage agent Pseudomonas lundensis ATCC 49968 by pyrimidines was examined. In P. lundensis cells grown on glucose as a carbon source, the enzymes aspartate transcarbamoylase, dihydroorotase, and orotidine 5′-monophosphate decarboxylase were induced by orotic acid. Pyrimidine auxotrophs containing reduced transcarbamoylase or orotate phosphoribosyltransferase activity were isolated using chemical mutagenesis and selection procedures. Independent of carbon source, the maximum derepression of enzyme activity was observed for orotidine 5′-monophosphate decarboxylase after pyrimidine limitation of either auxotroph. In the glucose-grown cells of the transcarbamoylase mutant strain, orotic acid induced dihydroorotase and decarboxylase activities. Aspartate transcarbamoylase activity in succinate-grown P. lundensis cells was highly regulated by pyrophosphate as well as by pyrimidine and purine ribonucleotides. It was concluded that pyrimidine formation in P. lundensis was controlled both at the level of de novo pyrimidine biosynthetic enzyme synthesis and at the level of transcarbamoylase activity.

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Nutrient utilization in actinomycetes. Induction of α-glucosidases in Streptomyces venezuelae

Canadian Journal of Microbiology ◽

10.1139/m81-098 ◽

1981 ◽

Vol 27 (7) ◽

pp. 639-645 ◽

Cited By ~ 27

Author(s):

S. Chatterjee ◽

L. C. Vining

Keyword(s):

Amino Acids ◽

Organic Acids ◽

Ribonucleic Acid ◽

De Novo ◽

Nutrient Utilization ◽

Enzyme Synthesis ◽

De Novo Synthesis ◽

Streptomyces Venezuelae ◽

Messenger Ribonucleic Acid ◽

Glucosidase Activity

Streptomyces venezuelae contains intracellular α-glucosidases that are induced during growth on maltose, isomaltose, maltotriose, dextrin, starch, and other α-glucosides. Induction was prevented by rifampicin at 10 μg∙mL−1 and inhibited by chloramphenicol or streptomycin, indicating that de novo synthesis of messenger ribonucleic acid and protein was required. Glucose and other readily utilizable sugars did not repress induction of α-glucosidase activity whereas certain organic acids and amino acids effectively reduced enzyme synthesis. Extracts of mycelium grown in the presence of maltose as an inducer hydrolysed maltose and isomaltose rapidly. Sucrose and other α-glucosides were less suitable substrates whereas trehalose and starch were not hydrolysed. No activity was observed with β-glucosides, α-galactosides, or methyl α-mannoside.

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Control of pyrimidine formation in Pseudomonas putida ATCC 17536

Canadian Journal of Microbiology ◽

10.1139/w02-110 ◽

2002 ◽

Vol 48 (12) ◽

pp. 1076-1081 ◽

Cited By ~ 12

Author(s):

Manuel F Santiago ◽

Thomas P West

Keyword(s):

Carbon Source ◽

Pseudomonas Putida ◽

Orotic Acid ◽

De Novo ◽

Enzyme Synthesis ◽

Pyrimidine Biosynthesis ◽

Transcriptional Level ◽

Orotate Phosphoribosyltransferase ◽

Biosynthesis Regulation ◽

Uracil Auxotrophs

The regulation of de novo pyrimidine biosynthesis in Pseudomonas putida ATCC 17536 by pyrimidines was explored. The pathway enzyme activities were higher in glucose-grown cells than in succinate-grown cells, indicating catabolite repression by succinate. In P. putida cells grown on succinate as a carbon source, only aspartate transcarbamoylase activity was greatly diminished by uracil supplementation. When glucose was the carbon source, orotic acid supplementation significantly decreased orotate phosphoribosyltransferase and orotidine 5'-monophosphate (OMP) decarboxylase activities. Uracil auxotrophs, deficient for dihydroorotase activity or with reduced phosphoribosyltransferase activity, were isolated. After pyrimidine limitation of both auxotrophs, the greatest derepression of enzyme activity was observed for OMP decarboxylase independent of carbon source. Orotic acid induced both phosphoribosyltransferase and decarboxylase activities in glucose-grown cells of the dihydroorotase-deficient strain. Regulation at the transcriptional level of de novo pyrimidine biosynthetic enzyme synthesis in P. putida ATCC 17536 was observed, which contrasts with previous observations.Key words: pyrimidine biosynthesis, regulation, auxotrophs, induction, Pseudomonas.

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Pichia stipitisL-rhamnose dehydrogenase and a catabolite-resistant mutant able to utilize 2-deoxy-D-glucose

Canadian Journal of Microbiology ◽

10.1139/m92-070 ◽

1992 ◽

Vol 38 (5) ◽

pp. 417-422 ◽

Cited By ~ 6

Author(s):

E. H. Pardo ◽

S. Funayama ◽

F. O. Pedrosa ◽

L. U. Rigo

Keyword(s):

Carbon Source ◽

Sole Carbon Source ◽

Glucose Repression ◽

Resistant Mutant ◽

Pichia Stipitis ◽

Enzyme Synthesis ◽

Selective Medium ◽

Wild Type ◽

Dehydrogenase Enzyme ◽

In The Wild

The yeast Pichia stipitis, strain NRC 5568, when grown on L-rhamnose as sole carbon source produced an NAD+-dependent L-rhamnose dehydrogenase enzyme, which is repressed by D-glucose. Mutants defective in carbon catabolite repression were isolated, using a selective medium containing 2-deoxy-D-glucose. Six of eight mutants resistant to 2-deoxy-D-glucose showed L-rhamnose dehydrogenase synthesis insensitive to D-glucose repression. All eight mutants, named PR mutants, as well as the parent strain, were found to grow on D-glucose, L-rhamnose, and glycerol. In addition, they were all capable of growing on 2-deoxy-D-glucose as sole carbon source, D-Glucose and 2-deoxy-D-glucose caused almost complete inhibition of L-rhamnose dehydrogenase synthesis in the wild-type strain but only a slight decrease in the enzyme synthesis in the mutant strain PR1. The wild-type and mutant strains showed the same pattern of inhibition by cycloheximide, 8-hydroxyquinoline, and benomyl. Key words: Pichia stipitis, L-rhamnose dehydrogenase, catabolite-resistant mutant, 2-deoxy-D-glucose.

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Characterization of Monocyte-Associated Factor V

Thrombosis and Haemostasis ◽

10.1055/s-0038-1649565 ◽

1993 ◽

Vol 70 (02) ◽

pp. 273-280 ◽

Cited By ~ 5

Author(s):

Janos Kappelmayer ◽

Satya P Kunapuli ◽

Edward G Wyshock ◽

Robert W Colman

Keyword(s):

Monoclonal Antibody ◽

Light Chain ◽

Peripheral Blood ◽

De Novo ◽

Factor V ◽

Blood Monocytes ◽

De Novo Synthesis ◽

Hep G2 ◽

Hep G2 Cells ◽

Peripheral Blood Monocytes

SummaryWe demonstrate that in addition to possessing binding sites for intact factor V (FV), unstimulated peripheral blood monocytes also express activated factor V (FVa) on their surfaces. FVa was identified on the monocyte surface by monoclonal antibody B38 recognizing FVa light chain and by human oligoclonal antibodies H1 (to FVa light chain) and H2 (to FVa heavy chain) using immunofluorescence microscopy and flow cytometry. On Western blots, partially cleaved FV could be identified as a 220 kDa band in lysates of monocytes. In addition to surface expression of FVa, monocytes also contain intracellular FV as detected only after permeabilization by Triton X-100 by monoclonal antibody B10 directed specifically to the Cl domain not present in FVa. We sought to determine whether the presence of FV in peripheral blood monocytes is a result of de novo synthesis.Using in situ hybridization, no FV mRNA could be detected in monocytes, while in parallel control studies, factor V mRNA was detectable in Hep G2 cells and CD18 mRNA in monocytes. In addition, using reverse transcriptase and the polymerase chain reaction, no FV mRNA was detected in mononuclear cells or in U937 cells, but mRNA for factor V was present in Hep G2 cells using the same techniques. These data suggest that FV is present in human monocytes, presumably acquired by binding of plasma FV, and that the presence of this critical coagulation factor is not due to de novo synthesis.

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Platelets Stimulate Thromboplastin Synthesis in Human Endothelial Cells

Thrombosis and Haemostasis ◽

10.1055/s-0038-1657323 ◽

1983 ◽

Vol 49 (02) ◽

pp. 069-072 ◽

Cited By ~ 20

Author(s):

U L H Johnsen ◽

T Lyberg ◽

K S Galdal ◽

H Prydz

Keyword(s):

Endothelial Cells ◽

De Novo ◽

Umbilical Vein ◽

Time Dependent ◽

De Novo Synthesis ◽

Human Endothelial Cells ◽

Tumor Promotor ◽

Coagulation Assay ◽

Platelet Aggregates ◽

Umbilical Vein Endothelial Cells

SummaryHuman umbilical vein endothelial cells in culture synthesize thromboplastin upon stimulation with phytohaemagglutinin (PHA) or the tumor promotor 12-O-tetradecanoyl-phorbol-13-acetate (TPA). The thromboplastin activity is further strongly enhanced in a time dependent reaction by the presence of gel-filtered platelets or platelet aggregates. This effect was demonstrable at platelet concentrations lower than those normally found in plasma, it may thus be of pathophysiological relevance. The thromboplastin activity increased with increasing number of platelets added. Cycloheximide inhibited the increase, suggesting that de novo synthesis of the protein component of thromboplastin, apoprotein III, is necessary.When care was taken to remove monocytes no thromboplastin activity and no apoprotein HI antigen could be demonstrated in suspensions of gel-filtered platelets, platelets aggregated with thrombin or homogenized platelets when studied with a coagulation assay and an antibody neutralization technique.

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