The membrane topology of vitamin K epoxide reductase is conserved between human isoforms and the bacterial enzyme

Cao, Z., Van Lith, M., Mitchell, L. J., Pringle, M. A. , Inaba, K. and Bulleid, N. J. (2016) The membrane topology of vitamin K epoxide reductase is conserved between human isoforms and the bacterial enzyme. Biochemical Journal, 473(7), pp. 851-858. (doi:10.1042/bj20151223) (PMID:26772871)

[img]
Preview
Text
120748.pdf - Accepted Version

600kB

Abstract

The membrane topology of vitamin K epoxide reductase (VKOR) is controversial with data supporting both a three transmembrane and a four transmembrane model. The positioning of the transmembrane domains and the loops between these domains is critical if we are to understand the mechanism of vitamin K oxidation and its recycling by members of the thioredoxin family of proteins and the mechanism of action of warfarin, an inhibitor of VKOR. Here we show that both mammalian VKOR isoforms adopt the same topology, with the large loop between transmembrane one and two facing the lumen of the endoplasmic reticulum (ER). We used a redox sensitive green fluorescent protein (GFP) fused to the N- or C-terminus to show that these regions face the cytosol, and introduction of glycosylation sites along with mixed disulfide formation with thioredoxin-like transmembrane protein (TMX) to demonstrate ER localization of the major loop. The topology is identical with the bacterial homologue from Synechococcus sp., for which the structure and mechanism of recycling has been characterized. Our results provide a resolution to the membrane topology controversy and support previous results suggesting a role for members of the ER protein disulfide isomerase (PDI) family in recycling VKOR.

Item Type:Articles
Status:Published
Refereed:Yes
Glasgow Author(s) Enlighten ID:Bulleid, Professor Neil and Van Lith, Dr Marcel and Cao, Dr Zhenbo and Pringle, Mrs Marie and Mitchell, Mrs Lorna
Authors: Cao, Z., Van Lith, M., Mitchell, L. J., Pringle, M. A., Inaba, K., and Bulleid, N. J.
College/School:College of Medical Veterinary and Life Sciences > Institute of Molecular Cell and Systems Biology
Journal Name:Biochemical Journal
Publisher:Portland Press Ltd.
ISSN:0264-6021
ISSN (Online):1470-8728
Published Online:15 January 2016
Copyright Holders:Copyright © 2016 The Authors
First Published:First published in Biochemical Journal 473(7):851-858
Publisher Policy:Reproduced in accordance with the copyright policy of the publisher

University Staff: Request a correction | Enlighten Editors: Update this record

Project CodeAward NoProject NamePrincipal InvestigatorFunder's NameFunder RefLead Dept
638691Identifying the reductive pathway in the mammalian endoplasmic reticulum.Neil BulleidBiotechnology and Biological Sciences Research Council (BBSRC)BB/L00593X/1RI MOLECULAR CELL & SYSTEMS BIOLOGY
652801Protein Folding and Thiol Modification in the Mammalian Endoplasmic ReticulumNeil BulleidWellcome Trust (WELLCOME)103720/Z/14/ZRI MOLECULAR CELL & SYSTEMS BIOLOGY