title: Identification and characterization of a cis,trans-mixed heptaprenyl diphosphate synthase from Arabidopsis thaliana.
abstract: In eukaryotes, dolichols (C(70-120)) play indispensable roles as glycosyl carrier lipids in the biosynthesis of glycoproteins on endoplasmic reticulum. In addition to dolichols, seed plants have other types of Z,E-mixed polyisoprenoids termed ficaprenol (tri-trans,poly-cis-polyprenol, C(45-75)) and betulaprenol (di-trans,poly-cis-polyprenol, C(30-45) and C(≥70)) in abundance. However, the physiological significance of these polyprenols has not been elucidated because of limited information regarding cis-prenyltransferases (cPTs) which catalyze the formation of the structural backbone of Z,E-mixed polyisoprenoids. In the comprehensive identification and characterization of cPT homologues from Arabidopsis thaliana, AtHEPS was identified as a novel cis,trans-mixed heptaprenyl diphosphate synthase. AtHEPS heterologously expressed in Escherichia coli catalyzed the formation of C(35) polyisoprenoid as a major product, independent of the chain lengths of all-trans allylic primer substrates. Kinetic analyses revealed that farnesyl diphosphate was the most favorable for AtHEPS among the allylic substrates tested suggesting that AtHEPS was responsible for the formation of C(35) betulaprenol. AtHEPS partially suppressed the phenotypes of a yeast cPT mutant deficient in the biosynthesis of dolichols. Moreover, in A. thaliana cells, subcellular localization of AtHEPS on the endoplasmic reticulum was shown by using green fluorescent protein fused proteins. However, a cold-stress-inducible expression of AtHEPS suggested that AtHEPS and its product might function in response to abiotic stresses rather than in cell maintenance as a glycosyl carrier lipid on the endoplasmic reticulum.
authors.affiliation: Department of Biomolecular Engineering, Graduate School of Engineering, Tohoku University, Sendai, Japan.
authors.forename: Kota
authors.initials: K
authors.lastname: Kera
authors.forename: Seiji
authors.initials: S
authors.lastname: Takahashi
authors.forename: Tsuyoshi
authors.initials: T
authors.lastname: Sutoh
authors.forename: Tanetoshi
authors.initials: T
authors.lastname: Koyama
authors.forename: Toru
authors.initials: T
authors.lastname: Nakayama
mesh_terms: D000040:Abscisic Acid / Q000494:pharmacology; D019883:Alkyl and Aryl Transferases / Q000145:classification / Q000235:genetics / Q000378:metabolism*; D017360:Arabidopsis / Q000166:cytology / Q000201:enzymology* / Q000235:genetics; D029681:Arabidopsis Proteins / Q000235:genetics / Q000378:metabolism*; D055162:Biocatalysis; D015153:Blotting, Western; D002413:Cations, Divalent / Q000494:pharmacology; D002478:Cells, Cultured; D003080:Cold Temperature; D004721:Endoplasmic Reticulum / Q000201:enzymology; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D018506:Gene Expression Regulation, Plant / Q000187:drug effects; D005816:Genetic Complementation Test; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D007700:Kinetics; D018613:Microscopy, Confocal; D009154:Mutation; D010802:Phylogeny; D010937:Plant Growth Regulators / Q000494:pharmacology; D011106:Polyisoprenyl Phosphates / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics / Q000254:growth & development; D012717:Sesquiterpenes / Q000378:metabolism; D013379:Substrate Specificity; D014166:Transferases / Q000145:classification / Q000235:genetics / Q000378:metabolism*
chemical_list: D029681:Arabidopsis Proteins; D002413:Cations, Divalent; D010937:Plant Growth Regulators; D011106:Polyisoprenyl Phosphates; D011994:Recombinant Proteins; D012717:Sesquiterpenes; D049452:Green Fluorescent Proteins; D000040:Abscisic Acid; C004808:farnesyl pyrophosphate; D014166:Transferases; D019883:Alkyl and Aryl Transferases; C577238:HEPS protein, Arabidopsis; C106821:cis-prenyl transferase; C024844:trans-hexaprenyltranstransferase