Purine metabolism in Saccharomyces cerevisiae

1977 ◽  
Vol 55 (9) ◽  
pp. 935-941 ◽  
Author(s):  
Philip W. Burridge ◽  
Robin A. Woods ◽  
J. Frank Henderson
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Nucleic Acid ◽  
De Novo ◽  
Adenine Nucleotides ◽  
Purine Metabolism ◽  
Guanine Nucleotides ◽  
Wild Type ◽  
Purine Bases

The synthesis, interconversion, and catabolism of purine bases, ribonucleosides, and ribonucleotides in wild-type Saccharomyces cerevisiae were studied by measuring the conversion of radioactive adenine, hypoxanthine, guanine, and glycine into acid-soluble purine bases, ribonucleosides, and ribonucleotides, and into nucleic acid adenine and guanine. The pathway(s) by which adenine is converted to inosinate is (are) uncertain. Guanine is extensively deaminated to xanthine. In addition, some guanine is converted to inosinate and adenine nucleotides. Inosinate formed either from hypoxanthine or de novo is readily converted to adenine and guanine nucleotides.

scholarly journals SPO71 Encodes a Developmental Stage-Specific Partner for Vps13 in Saccharomyces cerevisiae

2013 ◽  
Vol 12 (11) ◽  
pp. 1530-1537 ◽  
Author(s):  
Jae-Sook Park ◽  
Yuuya Okumura ◽  
Hiroyuki Tachikawa ◽  
Aaron M. Neiman
Keyword(s):  
Saccharomyces Cerevisiae ◽  
De Novo ◽  
Ectopic Expression ◽  
Membrane Vesicles ◽  
Specific Gene ◽  
Wild Type ◽  
Protein Mutation ◽  
Prospore Membrane ◽  
In The Wild ◽  
Membrane Morphogenesis

ABSTRACT The creation of haploid gametes in yeast, termed spores, requires the de novo formation of membranes within the cytoplasm. These membranes, called prospore membranes, enclose the daughter nuclei generated by meiosis. Proper growth and closure of prospore membranes require the highly conserved Vps13 protein. Mutation of SPO71 , a meiosis-specific gene first identified as defective in spore formation, was found to display defects in membrane morphogenesis very similar to those seen in vps13 Δ cells. Specifically, prospore membranes are smaller than in the wild type, they fail to close, and membrane vesicles are present within the prospore membrane lumen. As in vps13 Δ cells, the levels of phophatidylinositol-4-phosphate are reduced in the prospore membranes of spo71 Δ cells. SPO71 is required for the translocation of Vps13 from the endosome to the prospore membrane, and ectopic expression of SPO71 in vegetative cells results in mislocalization of Vps13. Finally, the two proteins can be coprecipitated from sporulating cells. We propose that Spo71 is a sporulation-specific partner for Vps13 and that they act in concert to regulate prospore membrane morphogenesis.

scholarly journals Improving 3-methylphenol (m-cresol) production in yeast via in vivo glycosylation or methylation

2020 ◽  
Vol 20 (8) ◽  
Author(s):  
Julia Hitschler ◽  
Eckhard Boles
Keyword(s):  
Saccharomyces Cerevisiae ◽  
De Novo ◽  
Wild Type ◽  
Biotechnological Production ◽  
Yeast Saccharomyces Cerevisiae ◽  
In Situ Extraction ◽  
Methylsalicylic Acid Synthase ◽  
In The Wild

ABSTRACT Heterologous expression of 6-methylsalicylic acid synthase (MSAS) together with 6-MSA decarboxylase enables de novo production of the platform chemical and antiseptic additive 3-methylphenol (3-MP) in the yeast Saccharomyces cerevisiae. However, toxicity of 3-MP prevents higher production levels. In this study, we evaluated in vivo detoxification strategies to overcome limitations of 3-MP production. An orcinol-O-methyltransferase from Chinese rose hybrids (OOMT2) was expressed in the 3-MP producing yeast strain to convert 3-MP to 3-methylanisole (3-MA). Together with in situ extraction by dodecane of the highly volatile 3-MA this resulted in up to 211 mg/L 3-MA (1.7 mM) accumulation. Expression of a UDP-glycosyltransferase (UGT72B27) from Vitis vinifera led to the synthesis of up to 533 mg/L 3-MP as glucoside (4.9 mM). Conversion of 3-MP to 3-MA and 3-MP glucoside was not complete. Finally, deletion of phosphoglucose isomerase PGI1 together with methylation or glycosylation and feeding a fructose/glucose mixture to redirect carbon fluxes resulted in strongly increased product titers, with up to 897 mg/L 3-MA/3-MP (9 mM) and 873 mg/L 3-MP/3-MP as glucoside (8.1 mM) compared to less than 313 mg/L (2.9 mM) product titers in the wild type controls. The results show that methylation or glycosylation are promising tools to overcome limitations in further enhancing the biotechnological production of 3-MP.

scholarly journals Purine salvage networks in Giardia lamblia.

1983 ◽  
Vol 158 (5) ◽  
pp. 1703-1712 ◽  
Author(s):  
C C Wang ◽  
S Aldritt
Keyword(s):  
Giardia Lamblia ◽  
De Novo ◽  
Adenine Nucleotides ◽  
Guanine Nucleotides ◽  
Rational Approach ◽  
Purine Nucleotides ◽  
Purine Salvage ◽  
Nucleotide Synthesis ◽  
Guanine Base

Purine metabolism in Giardia lamblia was investigated by monitoring incorporation of radiolabeled precursors into purine nucleotides in the log-phase trophozoites cultivated in vitro in axenic media and incubated in buffered saline glucose. The lack of incorporation of formate, glycine, hypoxanthine, inosine, and xanthine into the nucleotide pool suggests the absence of de novo purine nucleotide synthesis and the inability to form IMP as the precursor of AMP and GMP in G. lamblia. Only adenine, adenosine, guanine, and guanosine were incorporated. Further analysis of the labeled nucleotides by HPLC indicated that adenine and adenosine are converted only to adenine nucleotides whereas guanine and guanosine are only incorporated into guanine nucleotides. There is no competition of incorporation between adenine/adenosine and guanine/guanosine, and there is no interconversion between adenine and guanine nucleotides. Results from analyzing [5'-3H]guanosine incorporation indicate that the ribose moiety is not incorporated with the guanine base. Assays of purine salvage enzymic activities in the crude extracts of G. lamblia revealed the presence of only four major enzymes; adenosine and guanosine hydrolases and adenine and guanine phosphoribosyl transferases. Apparently, G. lamblia has an exceedingly simple purine salvage system; it converts adenosine and guanosine to corresponding purine bases and then forms AMP and GMP by the actions of corresponding purine phosphoribosyl transferases. The guanine phosphoribosyl transferase in G. lamblia is interesting because it does not recognize either hypoxanthine or xanthine as substrate. It thus must have a unique substrate specificity and may be regarded as a potential target to attack as a rational approach to chemotherapeutic control of giardiasis.

scholarly journals The effects of adenine nucleotides and guanine nucleotides on urate synthesis de novo by isolated chick liver and kidney cells

1975 ◽  
Vol 148 (3) ◽  
pp. 599-601 ◽  
Author(s):  
P Badenoch-Jones ◽  
P J Buttery
Keyword(s):  
De Novo ◽  
Adenine Nucleotides ◽  
Liver Cells ◽  
Guanine Nucleotides ◽  
Kidney Cells ◽  
Liver And Kidney

Isolated chick liver and kidney cells produce urate de novo from glycine, and this is partially inhibited by 1 mm-AMP and by 1 mm-GMP in liver cells but not in kidney cells. Azaserine fully inhibits this synthesis de novo, but attempts to isolate formylglycine amide ribonucleotide from azaserine-blocked cells were unsuccessful.

scholarly journals Saccharomyces cerevisiae Transcription Elongation Mutants Are Defective in PUR5 Induction in Response to Nucleotide Depletion

2000 ◽  
Vol 20 (20) ◽  
pp. 7427-7437 ◽  
Author(s):  
Randal J. Shaw ◽  
Daniel Reines
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Rna Polymerase Ii ◽  
De Novo ◽  
Transcription Elongation ◽  
Elongation Factor ◽  
Normal Growth ◽  
Luciferase Reporter ◽  
Wild Type ◽  
Luciferase Reporter Gene ◽  
Pol Ii

ABSTRACT IMP dehydrogenase (IMPDH) is the rate-limiting enzyme in the de novo synthesis of guanine nucleotides. It is a target of therapeutically useful drugs and is implicated in the regulation of cell growth rate. In the yeast Saccharomyces cerevisiae, mutations in components of the RNA polymerase II (Pol II) transcription elongation machinery confer increased sensitivity to a drug that inhibits IMPDH, 6-azauracil (6AU), by a mechanism that is poorly understood. This phenotype is thought to reflect the need for an optimally functioning transcription machinery under conditions of lowered intracellular GTP levels. Here we show that in response to the application of IMPDH inhibitors such as 6AU, wild-type yeast strains induce transcription of PUR5, one of four genes encoding IMPDH-related enzymes. Yeast elongation mutants sensitive to 6AU, such as those with a disrupted gene encoding elongation factor SII or those containing amino acid substitutions in Pol II subunits, are defective inPUR5 induction. The inability to fully inducePUR5 correlates with mutations that effect transcription elongation since 6AU-sensitive strains deleted for genes not related to transcription elongation are competent to induce PUR5. DNA encompassing the PUR5 promoter and 5′ untranslated region supports 6AU induction of a luciferase reporter gene in wild-type cells. Thus, yeast sense and respond to nucleotide depletion via a mechanism of transcriptional induction that restores nucleotides to levels required for normal growth. An optimally functioning elongation machinery is critical for this response.

scholarly journals Fitness effects ofcis-regulatory variants in theSaccharomyces cerevisiae TDH3promoter

2017 ◽  
Author(s):  
Fabien Duveau ◽  
William Toubiana ◽  
Patricia J. Wittkopp
Keyword(s):  
Gene Expression ◽  
Saccharomyces Cerevisiae ◽  
De Novo ◽  
Wild Type Allele ◽  
Wild Type ◽  
Fitness Effects ◽  
Regulatory Variants ◽  
Fitness Consequences ◽  
Regulatory Mutations ◽  
Affect Cell Growth

AbstractVariation in gene expression is widespread within and between species, but fitness consequences of this variation are generally unknown. Here we use mutations in theSaccharomyces cerevisiae TDH3promoter to assess how changes inTDH3expression affect cell growth. From these data, we predict the fitness consequences ofde novomutations and natural polymorphisms in theTDH3promoter. Nearly all mutations and polymorphisms in theTDH3promoter were found to have no significant effect on fitness in the environment assayed, suggesting that the wild type allele of this promoter is robust to the effects of most newcis-regulatory mutations.

scholarly journals Yeast peroxisomes multiply by growth and division

2007 ◽  
Vol 178 (3) ◽  
pp. 399-410 ◽  
Author(s):  
Alison M. Motley ◽  
Ewald H. Hettema
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Endoplasmic Reticulum ◽  
De Novo ◽  
Maturation Process ◽  
Wild Type ◽  
Related Proteins ◽  
Assay Conditions

Peroxisomes can arise de novo from the endoplasmic reticulum (ER) via a maturation process. Peroxisomes can also multiply by fission. We have investigated how these modes of multiplication contribute to peroxisome numbers in Saccharomyces cerevisiae and the role of the dynamin-related proteins (Drps) in these processes. We have developed pulse-chase and mating assays to follow the fate of existing peroxisomes, de novo–formed peroxisomes, and ER-derived preperoxisomal structures. We find that in wild-type (WT) cells, peroxisomes multiply by fission and do not form de novo. A marker for the maturation pathway, Pex3-GFP, is delivered from the ER to existing peroxisomes. Strikingly, cells lacking peroxisomes as a result of a segregation defect do form peroxisomes de novo. This process is slower than peroxisome multiplication in WT cells and is Drp independent. In contrast, peroxisome fission is Drp dependent. Our results show that peroxisomes multiply by growth and division under our assay conditions. We conclude that the ER to peroxisome pathway functions to supply existing peroxisomes with essential membrane constituents.

scholarly journals Physiological Properties of Saccharomyces cerevisiae from Which Hexokinase II Has Been Deleted

2001 ◽  
Vol 67 (4) ◽  
pp. 1587-1593 ◽  
Author(s):  
Jasper A. Diderich ◽  
Léonie M. Raamsdonk ◽  
Arthur L. Kruckeberg ◽  
Jan A. Berden ◽  
Karel Van Dam
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Glucose Repression ◽  
Adenine Nucleotides ◽  
Pyruvate Decarboxylase ◽  
Decarboxylase Activity ◽  
Diauxic Shift ◽  
Wild Type ◽  
Hexokinase Ii ◽  
In The Wild ◽  
Fructose 1,6 Bisphosphate

ABSTRACT Hexokinase II is an enzyme central to glucose metabolism and glucose repression in the yeast Saccharomyces cerevisiae. Deletion of HXK2, the gene which encodes hexokinase II, dramatically changed the physiology of S. cerevisiae. The hxk2-null mutant strain displayed fully oxidative growth at high glucose concentrations in early exponential batch cultures, resulting in an initial absence of fermentative products such as ethanol, a postponed and shortened diauxic shift, and higher biomass yields. Several intracellular changes were associated with the deletion of hexokinase II. Thehxk2 mutant had a higher mitochondrial H+-ATPase activity and a lower pyruvate decarboxylase activity, which coincided with an intracellular accumulation of pyruvate in the hxk2 mutant. The concentrations of adenine nucleotides, glucose-6-phosphate, and fructose-6-phosphate are comparable in the wild type and the hxk2 mutant. In contrast, the concentration of fructose-1,6-bisphosphate, an allosteric activator of pyruvate kinase, is clearly lower in the hxk2mutant than in the wild type. The results suggest a redirection of carbon flux in the hxk2 mutant to the production of biomass as a consequence of reduced glucose repression.

Arabidopsis 4-COUMAROYL-COA LIGASE 8 contributes to the biosynthesis of the benzenoid ring of coenzyme Q in peroxisomes

2019 ◽  
Vol 476 (22) ◽  
pp. 3521-3532
Author(s):  
Eric Soubeyrand ◽  
Megan Kelly ◽  
Shea A. Keene ◽  
Ann C. Bernert ◽  
Scott Latimer ◽  
...  
Keyword(s):  
Oxidative Metabolism ◽  
Time Course ◽  
Reverse Genetics ◽  
De Novo ◽  
Coenzyme Q ◽  
Control Level ◽  
Wild Type ◽  
Fluorescent Reporters ◽  
Coa Ligase ◽  
Peroxisomal Targeting

Plants have evolved the ability to derive the benzenoid moiety of the respiratory cofactor and antioxidant, ubiquinone (coenzyme Q), either from the β-oxidative metabolism of p-coumarate or from the peroxidative cleavage of kaempferol. Here, isotopic feeding assays, gene co-expression analysis and reverse genetics identified Arabidopsis 4-COUMARATE-COA LIGASE 8 (4-CL8; At5g38120) as a contributor to the β-oxidation of p-coumarate for ubiquinone biosynthesis. The enzyme is part of the same clade (V) of acyl-activating enzymes than At4g19010, a p-coumarate CoA ligase known to play a central role in the conversion of p-coumarate into 4-hydroxybenzoate. A 4-cl8 T-DNA knockout displayed a 20% decrease in ubiquinone content compared with wild-type plants, while 4-CL8 overexpression boosted ubiquinone content up to 150% of the control level. Similarly, the isotopic enrichment of ubiquinone's ring was decreased by 28% in the 4-cl8 knockout as compared with wild-type controls when Phe-[Ring-13C6] was fed to the plants. This metabolic blockage could be bypassed via the exogenous supply of 4-hydroxybenzoate, the product of p-coumarate β-oxidation. Arabidopsis 4-CL8 displays a canonical peroxisomal targeting sequence type 1, and confocal microscopy experiments using fused fluorescent reporters demonstrated that this enzyme is imported into peroxisomes. Time course feeding assays using Phe-[Ring-13C6] in a series of Arabidopsis single and double knockouts blocked in the β-oxidative metabolism of p-coumarate (4-cl8; at4g19010; at4g19010 × 4-cl8), flavonol biosynthesis (flavanone-3-hydroxylase), or both (at4g19010 × flavanone-3-hydroxylase) indicated that continuous high light treatments (500 µE m−2 s−1; 24 h) markedly stimulated the de novo biosynthesis of ubiquinone independently of kaempferol catabolism.

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