We have developed a purification procedure for the isolation of constitutive post-Golgi secretory vesicles from Saccharomyces cerevisiae. Although the post-Golgi stage of the secretion pathway is normally very rapid, we have used a temperature-sensitive secretory mutant, sec 6-4, to greatly expand the population of secretory vesicles. Following invertase as a marker, intact vesicles are enriched 36-fold from the crude lysate. The final preparation contains few contaminants as assessed by morphologic and biochemical examination. Three proteins (110, 40-45, and 18 kD) co-purify with the vesicle marker enzyme invertase. Metabolic labeling experiments indicate that these vesicle-associated proteins are synthesized during the period of vesicle accumulation. They are not apparent in the corresponding fractions from wild-type cells. Analysis of these proteins indicates that the 110-kD protein is a major glycoprotein residing in the vesicle lumen, while the 40-45- and 18-kD proteins are not glycosylated and are firmly associated with the vesicle membrane, each with at least one domain exposed on the cytoplasmic surface.
Article| July 01 1987
Purification and characterization of constitutive secretory vesicles from yeast.
N C Walworth
P J Novick
Online ISSN: 1540-8140
Print ISSN: 0021-9525
J Cell Biol (1987) 105 (1): 163–174.
N C Walworth, P J Novick; Purification and characterization of constitutive secretory vesicles from yeast.. J Cell Biol 1 July 1987; 105 (1): 163–174. doi: https://doi.org/10.1083/jcb.105.1.163
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