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The structure of FCGBP is formed as a disulfide-mediated homodimer between its C-terminal domains
Erik Ehrencrona1, Pablo Gallego1, Sergio Trillo-Muyo1
1Department of Medical Biochemistry and Cell Biology, Institute of Biomedicine, University of Gothenburg, Sweden.
Abstract:
Mucus in the colon is crucial for intestinal homeostasis by forming a barrier that separates microbes from the epithelium. This is achieved by the structural arrangement of the major mucus proteins, such as MUC2 and FCGBP, both of which are comprised of several von Willebrand D domains (vWD) and assemblies. Numerous disulfide bonds stabilise these domains, and intermolecular bonds generate multimers of MUC2. The oligomeric nature of FCGBP is not known. Human hFCGBP contains 13 vWD domains whereas mouse mFCGBP consists of only 7. We found unpaired cysteines in the vWD1 (human and mouse) and vWD5 (mouse)/vWD11 (human) assemblies which were not involved in disulfide bonds. However, the most C-terminal vWD domains, vWD7 (mouse)/vWD13 (human), formed disulfide-linked dimers. The intermolecular bond between C5284 and C5403 of human hFCGBP was observed by using mass spectrometry to generate the dimer. Cryo-EM structure analysis of recombinant mouse mFCGBP revealed a compact dimer with two symmetric intermolecular disulfide bonds between C2462 and C2581, corresponding to the dimerising cysteines in the human hFCGBP. This compact conformation involves interactions between the vWD assemblies, but although the domains involved at the interface are the same, the nature of the interactions differ. Mouse mFCGBP was also found to exist in a semi-extended conformation. These different interactions offer insights into the dynamic nature of the FCGBP homodimer.
Insights
The structure of the colon mucus protein FCGBP was investigated. Researchers discovered disulfide bonds in FCGBP that form dimers, revealing insights into its dynamic nature and role in intestinal homeostasis.
Area of Science:
- Biochemistry
- Structural Biology
- Gastroenterology
Background:
- Mucus in the colon is essential for intestinal homeostasis, forming a protective barrier.
- Major mucus proteins like MUC2 and FCGBP (Fibrinogen C-type lectin domain containing glycosaminoglycan binding protein) are stabilized by disulfide bonds.
- The oligomeric state and structural dynamics of FCGBP, a key mucus protein, were previously unknown.
Purpose of the Study:
- To elucidate the oligomeric structure and disulfide bond-mediated stabilization of human and mouse FCGBP.
- To understand the structural basis for FCGBP's role in maintaining the intestinal mucus barrier.
Main Methods:
- Mass spectrometry was used to identify disulfide bonds and characterize human FCGBP dimers.
- Cryo-electron microscopy (Cryo-EM) was employed to determine the structure of recombinant mouse FCGBP.
- Comparative analysis of human and mouse FCGBP structures to understand species-specific variations.
Main Results:
- Unpaired cysteines were identified in specific von Willebrand D (vWD) domains of both human and mouse FCGBP.
- The C-terminal vWD domains (vWD7 in mouse, vWD13 in human) form disulfide-linked dimers.
- Cryo-EM revealed a compact dimer structure for mouse FCGBP stabilized by two symmetric intermolecular disulfide bonds, with evidence for a semi-extended conformation.
Conclusions:
- FCGBP forms homodimers through specific intermolecular disulfide bonds in its C-terminal vWD domains.
- Structural variations and distinct interaction modes between human and mouse FCGBP highlight the dynamic nature of this homodimer.
- These findings provide crucial insights into the structural organization and function of FCGBP in the colonic mucus layer.
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