Isolation and identification of yeast plasma membrane

Schibeci, Angelo; Rattray, J. B. M.; Kidby, D. K.

doi:10.1016/0005-2736(73)90250-2

Cited by 61 publications

(30 citation statements)

References 26 publications

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“…The density of plasma membrane vesicles in sucrose gradients was between 1.16 and 1.18 g/cc (9, 10, 15-17, 30, 36, 37) which is similar to the density range reported for plasma membranes from cells of animals (5). The peak density of plasma membranes of algae (26,34), fungi, (1 1), and yeast (6,23,31) ranges from 1.18 to 1.23 g/cc.…”

supporting

confidence: 76%

Isolation of Plasma Membranes from Corn Roots by Sucrose Density Gradient Centrifugation

Leonard¹,

VanDerWoude²

1976

Plant Physiol.

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An investigation was conducted into the isolation of plasma membrane vesicles from primary roots of corn (Zea mays L., WF9 x M14) by sucrose density gradient centrifugation. Identification of plasma membranes in cell fractions was by specific staining with the periodic-chromic-phosphotungstic acid procedure. Plasma membrane vescles were rich in K+-stimulated ATPase activity at pH 6.5, and equilibrated in linear gradients of sucrose at a peak density of about 1.165 g/cc. It was necessary to remove mitochondria (equilibrium density of 1.18 g/cc) from the homogenate before density gradient centrifugation to minimize mitochondrial contamination of the plasma membrane fraction. Endoplasmic reticulum (NADH-cytochrome c reductase) and Golgi apparatus (latent IDPase) had equilibrium densities in sucrose of about 1.10 g/cc and 1.12 to 1.15 g/cc, respectively. A correlation (r = 0.975) was observed between K+-stimulated ATPase activity at pH 6.5 and the content of plasma membranes in various cell fractions. ATPase activity at pH 9 and cytochrome c oxidase activity were also correlated.A major peak of ATPase activity at pH 6.5 was observed at low density in Ficoll after nonequilibrium centrifugation in a combination Ficollsucrose gradient. Twenty to forty percent of the vesicles in this ATPase fraction stained positively for plasma membranes, and with equilibrium centrifugation the major portion of the ATPase activity shifted to densities in sucrose which were characteristic of plasma membranes. All major vesicular ATPase activities observed in Ficoll or sucrose contained substantial amounts of plasma membranes. For unknown reasons, mitochondria and plasma membranes equilibrated over a broader density range and at lower peak densities in sucrose as a result of equilibrium centrifugation through Ficoll.

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supporting

confidence: 76%

Isolation of Plasma Membranes from Corn Roots by Sucrose Density Gradient Centrifugation

Leonard¹,

VanDerWoude²

1976

Plant Physiol.

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“…In S. cerevisiae an oligomycin-insensitive ATPase has been shown to copurify with morphological markers also detectable in the plasma membrane of intact cells [2]. Phospholipids sterols and mannans [4-61 or nonpermeant radioactive tags of the plasma membrane [5] have also been shown to be present in a fraction enriched in oligomycin-insensitive ATPase activity. In S. pombe, we find that the distribution of mannans (but not that of cytochrome oxidase) parallels that of the pH-6.0 ATPase during differential and iso- [19,20] S. cerevisiae mannans have a triple localization : the endoplasmic reticulum, the so-called 'glucanase vesicles' and the plasma membrane.…”

Section: Discussionmentioning

confidence: 99%

Comparison of the Properties of Plasma Membrane‐Bound and Mitochondria‐Bound ATPases in the Yeast Schizosaccharomyces pombe

Dufour

Thines

et al. 1977

European Journal of Biochemistry

121

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Two distinct membrane‐bound ATPases of the glucose‐grown yeast Schizosaccharomyces pombe are separated by differential and isopycnic gradient centrifugation. The mitochondrial ATPase, with optimal activity at pH 9.0, equilibrates with cytochrome c oxidase at densities 1.17 and 1.19 g × cm−3 while the non‐mitochondrial ATPase, with optimal activity at pH 6.0 is associated with mannans in 1.21‐g × cm−3 particles. Cytochemical staining for carbohydrates confirms the plasma membrane origin of vesicles found in a subcellular fraction enriched about 10 times in pH‐6.0 ATPase activity. The following analogies are found between the two purified membrane‐bound ATPases. They hydrolyse preferentially ATP and require magnesium. They exhibit a Km of 3.3 mM for ATP‐Mg at pH 6.0. The two activities are inhibited by ADP, Dio‐9, dicyclohexylcarbodiimide, triethyltin, miconazole, sodium fluoride and p‐hydroxymercuribenzoate. Nevertheless, the plasma membrane‐bound ATPase is distinct from the mitochondria‐bound ATPase in the following properties. The plasma membrane‐bound ATPase is not sensitive to the antibody dressed against the purified mitochondrial enzyme. Excess of Mg2+ inhibits the mitochondria‐bound ATPase but not the plasma membrane one. The relative specificities for nucleoside triphosphates and cations of the two enzymes are different. In contrast to the mitochondrial enzyme, the plasma membrane‐bound ATPase is not stimulated by carbonate and maleate anions. The plasma membrane‐bound ATPase is not inhibited by the mitochondrial inhibitors oligomycin and venturicidin.

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“…In many of these reports the enzymes concerned have been assumed to occur in the plasma membrane and the isolation procedure has then been based on these assumptions. An alternative approach has been adopted by Schibeci, Rattray & Kidby (1973) and Marriott (1979, involving radioactive labelling of the plasma membrane of intact protoplasts. By following the distribution of radioactivity upon fractionation, Marriott (I 975) was able to obtain relatively pure preparations of plasma membranes from the yeast and mycelial forms of Candida albicans.…”

Section: Introductionmentioning

confidence: 99%

Enzymic Activity of Purified Plasma Membranes from the Yeast and Mycelial forms of Candida albicans

Marriott

1975

Journal of General Microbiology

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S U M M A R YPlasma membranes were isolated from the yeast and mycelial forms of Candida albicans as described previously (Marriott, 1975) and examined for the presence of several enzymes. Measurement of specific activities showed enrichment of Mg2+-dependent and Na+/K+-stimulated Mg2+-dependent adenosine triphosphatase and mannan synthetase, in the plasma membrane fractions from both morphological forms of the organism. However, acid and alkaline phosphatase, NADH oxidase and $-nucleotidase showed no such specific location.

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Isolation and identification of yeast plasma membrane

Cited by 61 publications

References 26 publications

Isolation of Plasma Membranes from Corn Roots by Sucrose Density Gradient Centrifugation

Isolation of Plasma Membranes from Corn Roots by Sucrose Density Gradient Centrifugation

Comparison of the Properties of Plasma Membrane‐Bound and Mitochondria‐Bound ATPases in the Yeast Schizosaccharomyces pombe

Enzymic Activity of Purified Plasma Membranes from the Yeast and Mycelial forms of Candida albicans

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