TWEAK [TNF (tumour necrosis factor)-like weak inducer of apoptosis] is a member of the TNF superfamily of cytokines. TWEAK binds with high affinity to a single TNF receptor super-family member, Fn14 (fibroblast growth factor-inducible 14). This interaction can stimulate a variety of biological responses, depending on the cell type analysed. The murine Fn14 extracellular region is only 53 amino acids in length and primarily consists of a CRD (cysteine-rich domain) containing three disulphide bonds. In the present study, we investigated whether TWEAK binding to this CRD was dependent on selected evolutionarily conserved amino acid residues by using a site-specific mutagenesis approach and several different ligand-binding assays. Our results indicate that three residues within the predicted Fn14 CRD A1 module (Asp45, Lys48 and Met50) and one residue within the predicted D2 module (Asp62) are each critical for high-affinity TWEAK binding. Mutation of the three charged polar residues Asp45, Lys48 and Asp62 had the greatest deleterious effect, suggesting that electrostatic interactions between TWEAK and Fn14 residues may be particularly important for complex formation or stability. To determine whether the four critical residues were likely to be located on the Fn14 CRD surface, we made an Fn14 homology model based on a previously derived X-ray structure for the B-cell maturation antigen receptor, which also contains only one CRD. This model revealed that each of these critical residues were in areas of the receptor that are potentially capable of interacting with TWEAK. These results indicate that the TWEAK–Fn14 interaction is highly dependent on multiple Fn14 residues located in both CRD modules.
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Research Article|
June 28 2006
TWEAK binding to the Fn14 cysteine-rich domain depends on charged residues located in both the A1 and D2 modules
Sharron A. N. Brown;
Sharron A. N. Brown
1Departments of Surgery and Physiology, Center for Vascular and Inflammatory Diseases, University of Maryland School of Medicine, 800 W. Baltimore St., Baltimore, MD 21201, U.S.A.
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Heather N. Hanscom;
Heather N. Hanscom
1Departments of Surgery and Physiology, Center for Vascular and Inflammatory Diseases, University of Maryland School of Medicine, 800 W. Baltimore St., Baltimore, MD 21201, U.S.A.
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Hong Vu;
Hong Vu
1
1Departments of Surgery and Physiology, Center for Vascular and Inflammatory Diseases, University of Maryland School of Medicine, 800 W. Baltimore St., Baltimore, MD 21201, U.S.A.
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Shelesa A. Brew;
Shelesa A. Brew
1Departments of Surgery and Physiology, Center for Vascular and Inflammatory Diseases, University of Maryland School of Medicine, 800 W. Baltimore St., Baltimore, MD 21201, U.S.A.
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Jeffrey A. Winkles
Jeffrey A. Winkles
2
1Departments of Surgery and Physiology, Center for Vascular and Inflammatory Diseases, University of Maryland School of Medicine, 800 W. Baltimore St., Baltimore, MD 21201, U.S.A.
2 To whom correspondence should be addressed (email jwinkles@som.umaryland.edu).
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Publisher: Portland Press Ltd
Received:
August 19 2005
Revision Received:
March 06 2006
Accepted:
March 09 2006
Accepted Manuscript online:
March 09 2006
Online ISSN: 1470-8728
Print ISSN: 0264-6021
The Biochemical Society, London
2006
Biochem J (2006) 397 (2): 297–304.
Article history
Received:
August 19 2005
Revision Received:
March 06 2006
Accepted:
March 09 2006
Accepted Manuscript online:
March 09 2006
Citation
Sharron A. N. Brown, Heather N. Hanscom, Hong Vu, Shelesa A. Brew, Jeffrey A. Winkles; TWEAK binding to the Fn14 cysteine-rich domain depends on charged residues located in both the A1 and D2 modules. Biochem J 15 July 2006; 397 (2): 297–304. doi: https://doi.org/10.1042/BJ20051362
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