TY - JOUR
T1 - Saccharomyces cerevisiae sec59 cells are deficient in dolichol kinase activity
AU - Heller, Loree
AU - Orlean, Peter
AU - Adair, W. Lee
PY - 1992
Y1 - 1992
N2 - The temperature-sensitive Saccharomyces cerevisiae mutant sec59 accumulates inactive and incompletely glycosylated protein precursors in its endoplasmic reticulum at the restrictive temperature. O-mannosylation and glycosyl phosphatidylinositol membrane anchoring of protein are also abolished, consistent with a deficiency in dolichyl phosphate mannose. Membranes prepared from sec59 cells that had been shifted to the restrictive temperature, however, made normal amounts of dolichyl phosphate mannose when exogenous dolichyl phosphate was supplied, but dolichyl phosphate mannose synthesis was severely depressed in the absence of exogenous dolichyl phosphate. Quantitative measurements of dolichyl phosphate in sec59 cells showed that the levels were decreased to 48% of wild type at the permissive temperature and to <10% at the restrictive temperature. Assays of enzymes from the dolichyl phosphate synthetic pathway, cis-prenyltransferase and dolichyl pyrophosphate phosphatase, gave wild-type levels. However, dolichol kinase activity was greatly decreased. When sec59 cells were transformed with a plasmid that over-expresses the wild-type gene, dolichol kinase activity increased 10-fold over wild-type levels. These results strongly suggest that the sec59 gene encodes dolichol kinase.
AB - The temperature-sensitive Saccharomyces cerevisiae mutant sec59 accumulates inactive and incompletely glycosylated protein precursors in its endoplasmic reticulum at the restrictive temperature. O-mannosylation and glycosyl phosphatidylinositol membrane anchoring of protein are also abolished, consistent with a deficiency in dolichyl phosphate mannose. Membranes prepared from sec59 cells that had been shifted to the restrictive temperature, however, made normal amounts of dolichyl phosphate mannose when exogenous dolichyl phosphate was supplied, but dolichyl phosphate mannose synthesis was severely depressed in the absence of exogenous dolichyl phosphate. Quantitative measurements of dolichyl phosphate in sec59 cells showed that the levels were decreased to 48% of wild type at the permissive temperature and to <10% at the restrictive temperature. Assays of enzymes from the dolichyl phosphate synthetic pathway, cis-prenyltransferase and dolichyl pyrophosphate phosphatase, gave wild-type levels. However, dolichol kinase activity was greatly decreased. When sec59 cells were transformed with a plasmid that over-expresses the wild-type gene, dolichol kinase activity increased 10-fold over wild-type levels. These results strongly suggest that the sec59 gene encodes dolichol kinase.
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U2 - 10.1073/pnas.89.15.7013
DO - 10.1073/pnas.89.15.7013
M3 - Article
C2 - 1323123
AN - SCOPUS:0026648113
VL - 89
SP - 7013
EP - 7016
JO - Proceedings of the National Academy of Sciences of the United States of America
JF - Proceedings of the National Academy of Sciences of the United States of America
SN - 0027-8424
IS - 15
ER -