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 e ssential for the assembly of a variety of glycoconjugates in the endoplasmi c reticulum of eukaryotic cells. In yeast, dolichols with chain lengths of 14-17 isoprene units are predominant, whereas in mammalian cells they conta in 19-22 isoprene units, In this biosynthetic pathway, t,t-farnesyl pyropho sphate is elongated to the appropriate long chain polyprenyl pyrophosphate by the sequential addition of cis-isoprene units donated by isopentenyl pyr ophosphate with t,t,c-geranylgeranyl pyrophosphate being the initial interm ediate formed. The condensation steps are catalyzed by cis-isoprenyltransfe rase (cis-IPTase), Genes encoding cis-IPTase activity have been identified in Micrococcus luteus, Escherichia coli, Arabidopsis thaliana, and Saccharo myces cerevisiae (RER2), Yeast cells deleted for the RER2 locus display a s evere 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 overexpr essing SRT1 have been determined by HPLC analysis. Dolichols and Dol-Ps wit h 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 SRTI was 7- to 17-fold higher than in microsomes from Delta rer2 cells. These results establish that yeast contains at leas t two cis-IPTases, and indicate that the chain length of dolichols is deter mined primarily by the enzyme catalyzing the chain elongation stage of the biosynthetic process.