Glycobiology, 2001, Vol. 11, No. 1 89-98
© 2001 Oxford University Press
An alternative cis-isoprenyltransferase activity in yeast that produces polyisoprenols with chain lengths similar to mammalian dolichols
Institute for Microbiology, ETH Zurich, CH-8092 Zurich, Switzerland, and 2Department of Biochemistry, University of Kentucky College of Medicine, Lexington, KY 40536, USA
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 (
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 rer2 cells overexpressing SRT1 was 7- to 17-fold higher than in microsomes from 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.
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