An alternative cis-isoprenyltransferase activity in yeast that produces polyisoprenols with chain lengths similar to mammalian dolichols.
Dolichyl monophosphate (Dol-P) is a polyisoprenoid glycosyl carrier lipid essential for the assembly of a variety of glycoconjugates in the endoplasmic reticulum of eukaryotic cells. In yeast, dolichols with chain lengths of 14--17 isoprene units are predominant, whereas in mammalian cells they contain 19--22 isoprene units. In this biosynthetic pathway, t,t-farnesyl pyrophosphate is elongated to the appropriate long chain polyprenyl pyrophosphate by the sequential addition of cis-isoprene units donated by isopentenyl pyrophosphate with t,t,c-geranylgeranyl pyrophosphate being the initial intermediate formed. The condensation steps are catalyzed by cis-isoprenyltransferase (cis-IPTase). Genes encoding cis-IPTase activity have been identified in Micrococcus luteus, Escherichia coli, Arabidopsis thaliana, and Saccharomyces cerevisiae (RER2). Yeast cells deleted for the RER2 locus display a severe growth defect, but are still viable, possibly due to the activity of an homologous locus, SRT1. The dolichol and Dol-P content of exponentially growing revertants of RER2 deleted cells (Delta rer2) and of cells overexpressing SRT1 have been determined by HPLC analysis. Dolichols and Dol-Ps with 19--22 isoprene units, unusually long for yeast, were found, and shown to be utilized for the biosynthesis of lipid intermediates involved in protein N-glycosylation. In addition, cis-IPTase activity in microsomes from Delta rer2 cells overexpressing SRT1 was 7- to 17-fold higher than in microsomes from Delta rer2 cells. These results establish that yeast contains at least two cis-IPTases, and indicate that the chain length of dolichols is determined primarily by the enzyme catalyzing the chain elongation stage of the biosynthetic process.
Schenk
B
B
Institute for Microbiology, ETH Zurich, CH-8092 Zurich, Switzerland.
Rush
J S
JS
Waechter
C J
CJ
Aebi
M
M
D019883:Alkyl and Aryl Transferases / Q000378:metabolism*; D000818:Animals; D001483:Base Sequence; D017931:DNA Primers; D004286:Dolichols / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism*; D006031:Glycosylation; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics; D017384:Sequence Deletion
D017931:DNA Primers; D004286:Dolichols; D019883:Alkyl and Aryl Transferases; C084921:cis-isoprenyltransferase