title: Hydrophilic anilinogeranyl diphosphate prenyl analogues are Ras function inhibitors.
abstract: Sequential processing of H-Ras by protein farnesyl transferase (FTase), Ras converting enzyme (Rce1), and protein-S-isoprenylcysteine O-methyltransferase (Icmt) to give H-Ras C-terminal farnesyl-S-cysteine methyl ester is required for appropriate H-Ras membrane localization and function, including activation of the mitogen-activated protein kinase (MAPK) cascade. We employed a Xenopus laevis oocyte whole-cell model system to examine whether anilinogeranyl diphosphate analogues of similar shape and size, but with a hydrophobicity different from that of the FTase substrate farnesyl diphosphate (FPP), could ablate biological function of H-Ras. Analysis of oocyte maturation kinetics following microinjection of in vitro analogue-modified H-Ras into isoprenoid-depleted oocytes revealed that analogues with a hydrophobicity near that of FPP supported H-Ras biological function, while the analogues p-nitroanilinogeranyl diphosphate (p-NO2-AGPP), p-cyanoanilinogeranyl diphosphate (p-CN-AGPP), and isoxazolaminogeranyl diphosphate (Isox-GPP) with hydrophobicities 2-5 orders of magnitude lower than that of FPP did not. We found that although H-Ras modified with FPP analogues p-NO2-AGPP, p-CN-AGPP, and Isox-GPP was an efficient substrate for C-terminal postprenylation processing by Rce1 and Icmt, co-injection of H-Ras with analogues p-NO2-AGPP, p-CN-AGPP, or Isox-GPP could not activate MAPK. We propose that H-Ras biological function requires a minimum lipophilicity of the prenyl group to allow important interactions downstream of the C-terminal processed H-Ras protein. The hydrophilic FPP analogues p-NO2-AGPP, p-CN-AGPP, and Isox-GPP are H-Ras function inhibitors (RFIs) and serve as lead compounds for a unique class of potential anticancer therapeutics.
authors.affiliation: Department of Molecular and Cellular Biochemistry, University of Kentucky, Lexington, Kentucky 40536-0084, USA.
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Roberts
authors.forename: Jerry M
authors.initials: JM
authors.lastname: Troutman
authors.forename: Kareem A H
authors.initials: KA
authors.lastname: Chehade
authors.forename: Hyuk C
authors.initials: HC
authors.lastname: Cha
authors.forename: Joseph P Y
authors.initials: JP
authors.lastname: Kao
authors.forename: Xiaoqin
authors.initials: X
authors.lastname: Huang
authors.forename: Chang-Guo
authors.initials: CG
authors.lastname: Zhan
authors.forename: Yuri K
authors.initials: YK
authors.lastname: Peterson
authors.forename: Thangaiah
authors.initials: T
authors.lastname: Subramanian
authors.forename: Srinivasan
authors.initials: S
authors.lastname: Kamalakkannan
authors.forename: Douglas A
authors.initials: DA
authors.lastname: Andres
authors.forename: H Peter
authors.initials: HP
authors.lastname: Spielmann
mesh_terms: D019883:Alkyl and Aryl Transferases / Q000378:metabolism; D000818:Animals; D002384:Catalysis; D010450:Endopeptidases / Q000378:metabolism; D057927:Hydrophobic and Hydrophilic Interactions; D008666:Metalloendopeptidases; D015689:Oncogene Protein p21(ras) / Q000037:antagonists & inhibitors* / Q000378:metabolism / Q000502:physiology*; D009865:Oocytes / Q000378:metabolism; D011106:Polyisoprenyl Phosphates / Q000138:chemical synthesis / Q000494:pharmacology*; D043484:Proprotein Convertases; D011496:Protein Methyltransferases / Q000378:metabolism; D017368:Protein Prenylation / Q000187:drug effects*; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism; D012717:Sesquiterpenes / Q000138:chemical synthesis / Q000494:pharmacology*; D014982:Xenopus laevis
chemical_list: D011106:Polyisoprenyl Phosphates; D029701:Saccharomyces cerevisiae Proteins; D012717:Sesquiterpenes; C004808:farnesyl pyrophosphate; D011496:Protein Methyltransferases; C056501:protein-S-isoprenylcysteine O-methyltransferase; D019883:Alkyl and Aryl Transferases; C064911:p21(ras) farnesyl-protein transferase; D010450:Endopeptidases; D043484:Proprotein Convertases; C105193:RCE1 protein, S cerevisiae; D008666:Metalloendopeptidases; D015689:Oncogene Protein p21(ras)