title: Disulfide-dependent protein folding is linked to operation of the vitamin K cycle in the endoplasmic reticulum. A protein disulfide isomerase-VKORC1 redox enzyme complex appears to be responsible for vitamin K1 2,3-epoxide reduction.
abstract: Gamma-carboxylation of vitamin K-dependent proteins is dependent on formation of reduced vitamin K1 (Vit.K1H2) in the endoplasmic reticulum (ER), where it works as an essential cofactor for gamma-carboxylase in post-translational gamma-carboxylation of vitamin K-dependent proteins. Vit.K1H2 is produced by the warfarin-sensitive enzyme vitamin K 2,3-epoxide reductase (VKOR) of the vitamin K cycle that has been shown to harbor a thioredoxin-like CXXC center involved in reduction of vitamin K1 2,3-epoxide (Vit.K>O). However, the cellular system providing electrons to the center is unknown. Here data are presented that demonstrate that reduction is linked to dithiol-dependent oxidative folding of proteins in the ER by protein disulfide isomerase (PDI). Oxidative folding of reduced RNase is shown to trigger reduction of Vit.K>O and gamma-carboxylation of the synthetic gamma-carboxylase peptide substrate FLEEL. In liver microsomes, reduced RNase-triggered gamma-carboxylation is inhibited by the PDI inhibitor bacitracin and also by small interfering RNA silencing of PDI in HEK 293 cells. Immunoprecipitation and two-dimensional SDS-PAGE of microsomal membrane proteins demonstrate the existence of a VKOR enzyme complex where PDI and VKORC1 appear to be tightly associated subunits. We propose that the PDI subunit of the complex provides electrons for reduction of the thioredoxin-like CXXC center in VKORC1. We can conclude that the energy required for gamma-carboxylation of proteins is provided by dithiol-dependent oxidative protein folding in the ER and thus is linked to de novo protein synthesis.
authors.affiliation: Department of Internal Medicine, Wake Forest University School of Medicine, Winston-Salem, North Carolina 27157, USA.
authors.forename: Nadeem
authors.initials: N
authors.lastname: Wajih
authors.forename: Susan M
authors.initials: SM
authors.lastname: Hutson
authors.forename: Reidar
authors.initials: R
authors.lastname: Wallin
mesh_terms: D000818:Animals; D001692:Biological Transport; D019735:Carbon-Carbon Ligases / Q000378:metabolism; D002460:Cell Line; D006224:Cricetinae; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D006899:Mixed Function Oxygenases / Q000378:metabolism*; D010084:Oxidation-Reduction; D019704:Protein Disulfide-Isomerases / Q000378:metabolism*; D017510:Protein Folding; D014050:Toluene / Q000031:analogs & derivatives; D014812:Vitamin K / Q000378:metabolism*; D010837:Vitamin K 1 / Q000031:analogs & derivatives* / Q000378:metabolism; D064417:Vitamin K Epoxide Reductases
chemical_list: D014812:Vitamin K; C016186:vitamin K1 oxide; D014050:Toluene; D010837:Vitamin K 1; D006899:Mixed Function Oxygenases; D064417:Vitamin K Epoxide Reductases; D019704:Protein Disulfide-Isomerases; D019735:Carbon-Carbon Ligases; C020660:glutamyl carboxylase; C004848:dithiol