Large scale isolation and storage of Neurospora plasma membranes

Stroobant, P.; Scarborough, G.A.

Analytical Biochemistry 95(2): 554-558

1979


ISSN/ISBN: 0003-2697
PMID: 156512
DOI: 10.1016/0003-2697(79)90771-1
Accession: 068522697

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Abstract
Functionally inverted plasma membrane vesicles isolated from the eukaryotic microorganism N. crassa generate and maintain a transmembrane electrical potential via ATP hydrolysis catalyzed by a plasma membrane ATPase. To facilitate investigation of the molecular mechanism of the electrogenic ATPase and other transport systems, a method was developed for the large-scale isolation and storage of Neurospora plasma membranes in a stable form. Large quantities of open plasma membrane sheets (ghosts) are isolated by a scaled-up modification of the original method and stored at -20.degree. C in 60% glycerol (vol/vol). As needed, the ghosts are washed free of glycerol and then converted to closed vesicles by a modification of the original method. With this technique, plasma membrane vesicles with normal electrogenic pump activity can be prepared daily in approximately 2.5 h.