A Rapid and Reproducible Method for Obtaining Yeast Protoplasts

Schwencke, Jaime; González, Guillermo; Farías, Gilda

doi:10.1002/j.2050-0416.1969.tb03176.x

Cited by 16 publications

(4 citation statements)

References 19 publications

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“…For the rapid conversion of cells to spheroplasts (1 5 -20 min) the pre-treatment with 100 mM Tris (pH 8), 5 mM EDTA and 5 mM dithiothreitol for 5 min at 30 "C [13] was found to be helpful. Spheroplasts were washed twice and finally resuspended into 0.6 M mannitol, 0.2 M KCl, 50 mM glucose and 50 mM 2-(N-morpholino)-ethane sulfonate adjusted to pH 6.0.…”

Section: Preparation Of Sp Her Oplas Tsmentioning

confidence: 99%

The Transport of S-Adenosyl-l-methionine in Isolated Yeast Vacuoles and Spheroplasts

Schwencke

Robichon-Szulmajstfr

1976

Eur J Biochem

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1. The properties of S-adenosyl-L-methionine accumulating system for both vacuoles and spheroplasts are described. Yeast vacuoles were obtained by a modified metabolic lysis procedure from spheroplasts of Saccharomyces cerevisiae.2. Isolated vacuoles accumulate S-adenosyl-L-methionine by means of a highly specific transport system as indicated by competition experiments with structural analogs of S-adenosyl-L-methionine. The S-adenosyl-L-methionine transport system shows saturation kinetics with an apparent K , of 68 pM in vacuoles and 11 pM in spheroplasts.3. S-Adenosyl-L-methionine accumulation into vacuoles does not require glucose, phosphoenolpyruvic acid, ATP, ADP nor any other tri-or di-phosphorylated nucleotides. It is insensitive to azide and 2,4-dinitrophenol which strongly inhibit the glucose-dependent accumulation of Sadenosyl-L-methionine in spheroplasts.4. The transport of S-adenosyl-L-methionine into vacuoles ic optimal at pH 7.4 and is insensitive to nystatin while the uptake of S-adenosyl-L-methionine into spheroplasts is optimal at pH 5.0 and is strongly sensitive to nystatin. On this basis it has thus been possible to measure both the intracytoplasmic and the intravacuolar pool of S-adenosyl-L-methionine.5. Our results indicate the existence of a highly specific S-adenosyl-L-methionine transport system in the vacuolar membrane which is clearly different from the one present in the plasma membrane of yeast cells.

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Section: Preparation Of Sp Her Oplas Tsmentioning

confidence: 99%

The Transport of S-Adenosyl-l-methionine in Isolated Yeast Vacuoles and Spheroplasts

Schwencke

Robichon-Szulmajstfr

1976

Eur J Biochem

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“…2 The lytic preparation.-The lytic enzyme preparation was obtained from a five-day culture of Strcptomyces WL-6 as reported.2 However, the precipitation with (NH^gSOŵ as performed in the presence of 1-mM EDTA and 0-5-mM 2-mercaptoethanol, a treatment which enhanced reproducibility.…”

Section: Methodsmentioning

confidence: 99%

General Applicability of a Method for Obtaining Yeast Protoplasts Using a Lytic Preparation From Streptomyces

Schwencke

Caglevic

Iäggli

1971

Journal of the Institute of Brewing

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The ability of a lytic preparation from Streptomyces WL‐6 to produce yeast protoplasts was analysed. True protoplasts were found to be produced from strains of the following yeasts: Trigonopsis variabilis, Toruiopsis stellata, Candida puicherrima, Saccharomyces carisbergensis, Saccharomyces cerevisiae and Saccharomyces meilis. The time necessary to obtain 100% protoplasts varied from 35 to 50 min.

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“…18 ports (2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13)(14)(15)(16)(17) showing that various microorganisms accumulate enzymes in their culture media with lytic activities against yeast cell walls. Some of these, however, only lyse yeast cell walls effectively after the yeast has been subjected to heat or acetone treatment ( 3,5,8,9,11,16).…”

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confidence: 99%

“…Some of these, however, only lyse yeast cell walls effectively after the yeast has been subjected to heat or acetone treatment ( 3,5,8,9,11,16). Further, some enzymes only lyse viable yeast cells at certain growth stages ( 4,7,12,13,15) andjor after treatment of the cells with 2-mercaptoethanol or dithiothreitol (10,13,15). Recently,MADA et al (17) tested more than 90 strains of Basidiomycetes to obtain enzymes that lyse viable yeast cells, and reported that one of these strains could lyse viable cells of Saccharomyces cerevisiae without any preliminary treatment of the cells.…”

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confidence: 99%

Lysis of Viable Yeast Cells by Enzymes of Arthrobacter Luteus

Kitamura

Kaneko

Yamämoto

1972

J. Gen. Appl. Microbiol.

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Some organisms capable of lysing viable yeast cells were isolated from brewery sewage by the enrichment culture method by intermittent feeding with brewer's yeast. The strain (B 111-1) showing the highest lytic activity was identified as Arthrobacter luteus.The activity of Arthrobacter luteus, B 111-1, to lyse viable yeast cells appeared in the culture medium in the early stationary phase of the bacterial growth.The pattern of change in lytic activity during cultivation was different from those of IS-1, 3-glucanase and protease activities suggesting that some additional factors are essential for the lysis of viable yeast cells.The pH value of culture medium during cultivation influenced the development of the activity considerably.The pH value first decreased and then increased with the appearance of lytic activity.The activity to lyse viable yeast cells developed adaptively when the organism was cultivated in a medium containing yeast cells or 3-1, 3-glucan, suggesting that the latter causes induction.Cell-free culture fluid of the organism showed a lytic activity-on cells of brewer's yeast at all stages of growth and on viable cells of yeast strains belonging to a wide range of genera. However, it did not lyse viable cells of the yeasts Rhodoturula, Pichia, and Hansenula, or viable and heat-treated cells of bacteria and molds.The lytic activity of the cell-free culture fluid was not lost by dialysis. It was salted out at 0.395 saturation of ammonium sulfate and was lost on incubation at 60° for 5 min. It was also inactivated by such protein denaturing agents as mercuric chloride, sodium laurylbenzene sulfonate, and silver nitrate, indicating that the lytic activity was due to enzymes.The optimum pH for lysis of viable yeast cells was 7.0-8.0. The lytic activity was relatively stable between pH 5 and 10.Since GIAJA's work (1) on snail gut juice, there have been 57 many re-

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A Rapid and Reproducible Method for Obtaining Yeast Protoplasts

Cited by 16 publications

References 19 publications

The Transport of S-Adenosyl-l-methionine in Isolated Yeast Vacuoles and Spheroplasts

The Transport of S-Adenosyl-l-methionine in Isolated Yeast Vacuoles and Spheroplasts

General Applicability of a Method for Obtaining Yeast Protoplasts Using a Lytic Preparation From Streptomyces

Lysis of Viable Yeast Cells by Enzymes of Arthrobacter Luteus

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