HJV (haemojuvelin) plays a key role in iron metabolism in mammals by regulating expression of the liver-derived hormone hepcidin, which controls systemic iron uptake and release. Mutations in HJV cause juvenile haemochromatosis, a rapidly progressing iron overload disorder in humans. HJV, also known as RGMc (repulsive guidance molecule c), is a member of the three-protein RGM family. RGMs are GPI (glycosylphosphatidylinositol)-linked glycoproteins that share ~50% amino acid identity and several structural motifs, including the presence of 14 cysteine residues in analogous locations. Unlike RGMa and RGMb, HJV/RGMc is composed of both single-chain and two-chain isoforms. To date there is no structural information for any member of the RGM family. In the present study we have mapped the disulfide bonds in mouse HJV/RGMc using a proteomics strategy combining sequential MS steps composed of ETD (electron transfer dissociation) and CID (collision-induced dissociation), in which ETD induces cleavage of disulfide linkages, and CID establishes disulfide bond assignments between liberated peptides. The results of the present study identified an HJV/RGMc molecular species containing four disulfide linkages. We predict using ab initio modelling that this molecule is a single-chain HJV/RGMc isoform. Our observations outline a general approach using tandem MS and ab initio molecular modelling to define unknown structural features in proteins.
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Research Article|
April 25 2013
Proteomic analysis and molecular modelling characterize the iron-regulatory protein haemojuvelin/repulsive guidance molecule c
Mahta Nili;
Mahta Nili
1Department of Biochemistry and Molecular Biology, Oregon Health and Science University, Portland, OR 97239-3098, U.S.A.
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Larry David;
Larry David
1Department of Biochemistry and Molecular Biology, Oregon Health and Science University, Portland, OR 97239-3098, U.S.A.
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Johannes Elferich;
Johannes Elferich
1Department of Biochemistry and Molecular Biology, Oregon Health and Science University, Portland, OR 97239-3098, U.S.A.
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Ujwal Shinde;
Ujwal Shinde
1Department of Biochemistry and Molecular Biology, Oregon Health and Science University, Portland, OR 97239-3098, U.S.A.
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Peter Rotwein
Peter Rotwein
1
1Department of Biochemistry and Molecular Biology, Oregon Health and Science University, Portland, OR 97239-3098, U.S.A.
1To whom correspondence should be addressed (email rotweinp@ohsu.edu).
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Publisher: Portland Press Ltd
Received:
December 07 2012
Revision Received:
February 18 2013
Accepted:
March 06 2013
Accepted Manuscript online:
March 06 2013
Online ISSN: 1470-8728
Print ISSN: 0264-6021
© The Authors Journal compilation © 2013 Biochemical Society
2013
Biochem J (2013) 452 (1): 87–95.
Article history
Received:
December 07 2012
Revision Received:
February 18 2013
Accepted:
March 06 2013
Accepted Manuscript online:
March 06 2013
Citation
Mahta Nili, Larry David, Johannes Elferich, Ujwal Shinde, Peter Rotwein; Proteomic analysis and molecular modelling characterize the iron-regulatory protein haemojuvelin/repulsive guidance molecule c. Biochem J 15 May 2013; 452 (1): 87–95. doi: https://doi.org/10.1042/BJ20121845
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