scholarly journals The effect of pre-irradiation culture conditions on the yield of X-ray-induced respiratory-deficient mutants in haploid Saccharomyces cerevisiae*

Hereditas ◽  
2009 ◽  
Vol 95 (2) ◽  
pp. 333-335
Author(s):  
TRYGVE EKLUND
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Culture Conditions ◽  
X Ray ◽  
Respiratory Deficient

scholarly journals Mutation of High-Affinity Methionine Permease Contributes to Selenomethionyl Protein Production in Saccharomyces cerevisiae

2010 ◽  
Vol 76 (19) ◽  
pp. 6351-6359 ◽  
Author(s):  
Toshihiko Kitajima ◽  
Yasunori Chiba ◽  
Yoshifumi Jigami
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Methylotrophic Yeast ◽  
Culture Conditions ◽  
Crystallographic Analysis ◽  
Gene Encoding ◽  
X Ray ◽  
X Ray Crystallography ◽  
High Affinity ◽  
Standard Culture ◽  
Resistant Mutants

ABSTRACT The production of selenomethionine (SeMet) derivatives of recombinant proteins allows phase determination by single-wavelength or multiwavelength anomalous dispersion phasing in X-ray crystallography, and this popular approach has permitted the crystal structures of numerous proteins to be determined. Although yeast is an ideal host for the production of large amounts of eukaryotic proteins that require posttranslational modification, the toxic effects of SeMet often interfere with the preparation of protein derivatives containing this compound. We previously isolated a mutant strain (SMR-94) of the methylotrophic yeast Pichia pastoris that is resistant to both SeMet and selenate and demonstrated its applicability for the production of proteins suitable for X-ray crystallographic analysis. However, the molecular basis for resistance to SeMet by the SMR-94 strain remains unclear. Here, we report the characterization of SeMet-resistant mutants of Saccharomyces cerevisiae and the identification of a mutant allele of the MUP1 gene encoding high-affinity methionine permease, which confers SeMet resistance. Although the total methionine uptake by the mup1 mutant (the SRY5-7 strain) decreased to 47% of the wild-type level, it was able to incorporate SeMet into the overexpressed epidermal growth factor peptide with 73% occupancy, indicating the importance of the moderate uptake of SeMet by amino acid permeases other than Mup1p for the alleviation of SeMet toxicity. In addition, under standard culture conditions, the mup1 mutant showed higher productivity of the SeMet derivative relative to other SeMet-resistant mutants. Based on these results, we conclude that the mup1 mutant would be useful for the preparation of selenomethionyl proteins for X-ray crystallography.

Effects of elevated temperature on growth, respiratory-deficient mutation, respiratory activity, and ethanol production in yeast

1988 ◽  
Vol 34 (8) ◽  
pp. 1014-1017 ◽  
Author(s):  
Midori Yamamura ◽  
Yoichi Nagami ◽  
Vitchuporn Vongsuvanlert ◽  
Jaroon Kumnuanta ◽  
Teijiro Kamihara
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Elevated Temperature ◽  
Yeast Extract ◽  
Respiratory Activity ◽  
Culture Conditions ◽  
Inoculum Size ◽  
Yeast Strains ◽  
Thermotolerant Strain ◽  
Grown Culture ◽  
Respiratory Deficient

Some mesophilic yeasts and a thermotolerant strain of Saccharomyces cerevisiae were found to grow at 40 °C in complex media containing 1% yeast extract when an inoculum of 106 or more cells∙mL−1 was used. Yeast extract (6%) permitted Saccharomyces cerevisiae to grow at 40 °C even with a smaller inoculum size (105 cells∙mL−1). The fraction of respiratory-deficient (petite) mutants in 40 °C grown culture was less than 10% except for the thermotolerant strain, which showed greatly increased levels depending on culture conditions. Seven of eight yeast strains exhibited extremely reduced cytochrome oxidase activity when grown at 40 °C irrespective of the frequency of the petite mutation. In contrast, the accumulation of ethanol in the medium and the ethanol-producing activity of the cells were not affected by growth at 40 °C.

scholarly journals INCREASED SPONTANEOUS MITOTIC SEGREGATION IN MMS-SENSITIVE MUTANTS OF SACCHAROMYCES CEREVISIAE

Genetics ◽  
1977 ◽  
Vol 87 (2) ◽  
pp. 229-236
Author(s):  
Satya Prakash ◽  
Louise Prakash
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Methyl Methanesulfonate ◽  
X Rays ◽  
X Ray ◽  
Complementation Groups

ABSTRACT Methyl methanesulfonate (MMS)-sensitive mutants of Saccharomyces cerevisiae belonging to four different complementation groups, when homozygous, increase the rate of spontaneous mitotic segregation to canavanine resistance from heterozygous sensitive (canr/+) diploids by 13- to 170-fold. The mms8—1 mutant is MMS and X-ray sensitive and increases the rate of spontaneous mitotic segregation 170-fold. The mms9—1 and mms13—1 mutants are sensitive to X rays and UV, respectively, in addition to MMS, and increase the rate of spontaneous mitotic segregation by 13-fold and 85-fold, respectively. The mutant mms21—1 is sensitive to MMS, X rays and UV and increases the rate of spontaneous mitotic segregation 23-fold.

scholarly journals Mobilization of Selenite by Ralstonia metallidurans CH34

2001 ◽  
Vol 67 (2) ◽  
pp. 769-773 ◽  
Author(s):  
Murielle Roux ◽  
Géraldine Sarret ◽  
Isabelle Pignot-Paintrand ◽  
Marc Fontecave ◽  
Jacques Coves
Keyword(s):  
Alcaligenes Eutrophus ◽  
Culture Conditions ◽  
Lag Phase ◽  
Elemental Selenium ◽  
Kinetics Analysis ◽  
X Ray ◽  
Polluted Sites ◽  
Ralstonia Metallidurans ◽  
X Ray Absorption ◽  
Bioremediation Processes

ABSTRACT Ralstonia metallidurans CH34 (formerlyAlcaligenes eutrophus CH34) is a soil bacterium characteristic of metal-contaminated biotopes, as it is able to grow in the presence of a variety of heavy metals. R. metalliduransCH34 is reported now to resist up to 6 mM selenite and to reduce selenite to elemental red selenium as shown by extended X-ray absorption fine-structure analysis. Growth kinetics analysis suggests an adaptation of the cells to the selenite stress during the lag-phase period. Depending on the culture conditions, the medium can be completely depleted of selenite. Selenium accumulates essentially in the cytoplasm as judged from electron microscopy and energy-dispersive X-ray analysis. Elemental selenium, highly insoluble, represents a nontoxic storage form for the bacterium. The ability of R. metallidurans CH34 to reduce large amounts of selenite may be of interest for bioremediation processes targeting selenite-polluted sites.

scholarly journals The Effects of Culture Conditions on the Secretion of Human Lysozyme by Saccharomyces cerevisiae A2-1-1A

1989 ◽  
Vol 53 (10) ◽  
pp. 2687-2694
Author(s):  
Kimihisa Ichikawa ◽  
Koji Komiya ◽  
Katsunori Suzuki ◽  
Tadaatsu Nakahara ◽  
Yoshifumi Jigami
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Culture Conditions ◽  
Human Lysozyme

scholarly journals Optimization of Saccharomyces cerevisiae α-galactosidase production and application in the degradation of raffinose family oligosaccharides

2019 ◽  
Vol 18 (1) ◽  
Author(s):  
María-Efigenia Álvarez-Cao ◽  
María-Esperanza Cerdán ◽  
María-Isabel González-Siso ◽  
Manuel Becerra
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Food Industry ◽  
Culture Conditions ◽  
Raffinose Family Oligosaccharides ◽  
Recombinant Production ◽  
Gene Coding ◽  
Productive Process ◽  
Hydrolysis Of ◽  
Degree Of Purification

Abstract Background α-Galactosidases are enzymes that act on galactosides present in many vegetables, mainly legumes and cereals, have growing importance with respect to our diet. For this reason, the use of their catalytic activity is of great interest in numerous biotechnological applications, especially those in the food industry directed to the degradation of oligosaccharides derived from raffinose. The aim of this work has been to optimize the recombinant production and further characterization of α-galactosidase of Saccharomyces cerevisiae. Results The MEL1 gene coding for the α-galactosidase of S. cerevisiae (ScAGal) was cloned and expressed in the S. cerevisiae strain BJ3505. Different constructions were designed to obtain the degree of purification necessary for enzymatic characterization and to improve the productive process of the enzyme. ScAGal has greater specificity for the synthetic substrate p-nitrophenyl-α-d-galactopyranoside than for natural substrates, followed by the natural glycosides, melibiose, raffinose and stachyose; it only acts on locust bean gum after prior treatment with β-mannosidase. Furthermore, this enzyme strongly resists proteases, and shows remarkable activation in their presence. Hydrolysis of galactose bonds linked to terminal non-reducing mannose residues of synthetic galactomannan-oligosaccharides confirms that ScAGal belongs to the first group of α-galactosidases, according to substrate specificity. Optimization of culture conditions by the statistical model of Response Surface helped to improve the productivity by up to tenfold when the concentration of the carbon source and the aeration of the culture medium was increased, and up to 20 times to extend the cultivation time to 216 h. Conclusions ScAGal characteristics and improvement in productivity that have been achieved contribute in making ScAGal a good candidate for application in the elimination of raffinose family oligosaccharides found in many products of the food industry.

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