| 1. |
- Ehrencrona, Erik, et al.
(författare)
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The IgGFc-binding protein FCGBP is secreted with all GDPH sequences cleaved but maintained by interfragment disulfide bonds
- 2021
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Ingår i: Journal of Biological Chemistry. - : Elsevier BV. - 0021-9258. ; 297:1
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Tidskriftsartikel (refereegranskat)abstract
- Mucus forms an important protective barrier that minimizes bacterial contact with the colonic epithelium. Intestinal mucus is organized in a complex network with several specific proteins, including the mucin-2 (MUC2) and the abundant IgGFc-binding protein, FCGBP. FCGBP is expressed in all intestinal goblet cells and is secreted into the mucus. It is comprised of repeated von Willebrand D (vWD) domain assemblies, most of which have a GDPH amino acid sequence that can be autocatalytically cleaved, as previously observed in the mucins MUC2 and mucin-5AC. However, the functions of FCGBP in the mucus are not understood. We show that all vWD domains of FCGBP with a GDPH sequence are cleaved and that these cleavages occur early during biosynthesis in the endoplasmic reticulum. All cleaved fragments, however, remain connected via a disulfide bond within each vWD domain. This cleavage generates a C-terminal-reactive Asp-anhydride that could react with other molecules, such as MUC2, but this was not observed. Quantitative analyses by MS showed that FCGBP was mainly soluble in chaotropic solutions, whereas MUC2 was insoluble, and most of the secreted FCGBP was not covalently bound to MUC2. Although FCGBP has been suggested to bind immunoglobulin G, we were unable to reproduce this binding in vitro using purified proteins. In conclusion, while the function of FCGBP is still unknown, our results suggest that it does not contribute to covalent crosslinking in the mucus, nor incorporate immunoglobulin G into mucus, instead the single disulfide bond linking each fragment could mediate controlled dissociation.
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| 2. |
- Ermund, Anna, et al.
(författare)
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Assembly, Release, and Transport of Airway Mucins in Pigs and Humans
- 2018
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Ingår i: Annals of the American Thoracic Society. - 1546-3222. ; 15
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Tidskriftsartikel (refereegranskat)abstract
- The respiratory system is protected from inhaled particles and microbes by the mucociliary system. This system differs between animal species, where pigs and humans have numerous submucosal glands. The polymer-forming mucin, MUC5B, is packed in a highly organized way in granules of the mucus-secreting cells in the glands. Upon secretion, the packed MUC5B is flushed out by a chloride-and bicarbonate-rich fluid from the cystic fibrosis transmembrane conductance regulator-expressing serosal cells located at the most distal part of the gland. The bicarbonate raises the pH and removes calcium from the N terminus of MUC5B, allowing the mucin to be pulled out into a linear polymer. Thousands of such polymers gather in bundles in the submucosal gland duct, and these bundles appear at the opening of the glands. They are moved by the beating cilia, and sweep over the airway surface and are patchily coated with the MUC5AC mucin from the surface goblet cells. The movement of these bundles is controlled by the MUC5AC mucin attachment/detachment to the goblet cells. Thus, higher animals with submucosal glands and large diameters of the proximal airways are efficiently cleaned by the thick mucus bundles sweeping the airway surface and moving particles and bacteria toward the larynx.
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| 3. |
- Ermund, Anna, et al.
(författare)
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The normal trachea is cleaned by MUC5B mucin bundles from the submucosal glands coated with the MUC5AC mucin
- 2017
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Ingår i: Biochemical and Biophysical Research Communications - BBRC. - : Elsevier BV. - 0006-291X .- 1090-2104. ; 492:3, s. 331-337
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Tidskriftsartikel (refereegranskat)abstract
- To understand the mucociliary clearance system, mucins were visualized by light, confocal and electron microscopy, and mucus was stained by Alcian blue and tracked by video microscopy on tracheal explants of newborn piglets. We observed long linear mucus bundles that appeared at the submucosal gland openings and were transported cephalically. The mucus bundles were shown by mass spectrometry and immunostaining to have a core made of MUC5B mucin and were coated with MUC5AC mucin produced by surface goblet cells. The transport speed of the bundles was slower than the airway surface liquid flow. We suggest that the goblet cell MUC5AC mucin anchors the mucus bundles and thus controls their transport. Normal clearance of the respiratory tree of pigs and humans, both rich in submucosal glands, is performed by thick and long mucus bundles. (C) 2017 The Authors. Published by Elsevier Inc.
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| 4. |
- Fakih, Dalia, et al.
(författare)
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Normal murine respiratory tract has its mucus concentrated in clouds based on the Muc5b mucin
- 2020
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Ingår i: American Journal of Physiology-Lung Cellular and Molecular Physiology. - : American Physiological Society. - 1040-0605 .- 1522-1504. ; 318:6
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Tidskriftsartikel (refereegranskat)abstract
- The organization of the normal airway mucus system differs in small experimental animals from that in humans and large mammals. To address normal murine airway mucociliary clearance, Alcian blue-stained mucus transport was measured ex vivo on tracheal tissues of naive C57BL/6, Muc5b(-/-), Muc5ac(-/-), and EGFP-tagged Muc5b reporter mice. Close to the larynx with a few submucosal glands, the mucus appeared as thick bundles. More distally in the trachea and in large bronchi, Alcian blue-stained mucus was organized in cloud-like formations based on the Muc5b mucin. On tilted tissue, the mucus clouds moved upward toward the larynx with an average velocity of 12 mu m/s compared with 20 mu m/s for beads not associated with clouds. In Muc5ac(-/-) mice, Muc5b formed mucus strands attached to the tissue surface, while in Muc5b(-/-) mice, Muc5ac had a more variable appearance. The normal mouse lung mucus thus appears as discontinuous clouds, clearly different from the stagnant mucus layer in diseased lungs.
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| 5. |
- Gallego, Pablo, et al.
(författare)
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The intestinal MUC2 mucin C-terminus is stabilized by an extra disulfide bond in comparison to von Willebrand factor and other gel-forming mucins
- 2023
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Ingår i: Nature communications. - 2041-1723. ; 14
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Tidskriftsartikel (refereegranskat)abstract
- The MUC2 mucin polymer is the main building unit of the intestinal mucus layers separating intestinal microbiota from the host epithelium. The MUC2 mucin is a large glycoprotein with a C-terminal domain similar to the MUC5AC and MUC5B mucins and the von Willebrand factor (VWF). A structural model of the C-terminal part of MUC2, MUC2-C, was generated by combining Cryo-electron microscopy, AlphaFold prediction, information of its glycosylation, and small angle X-ray scattering information. The globular VWD4 assembly in the N-terminal of MUC2-C is followed by 3.5 linear VWC domains that form an extended flexible structure before the C-terminal cystine-knot. All gel-forming mucins and VWF form tail-tail disulfide-bonded dimers in their C-terminal cystine-knot domain, but interestingly the MUC2 mucin has an extra stabilizing disulfide bond on the N-terminal side of the VWD4 domain, likely essential for a stable intestinal mucus barrier.
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| 6. |
- Recktenwald, Christian V, et al.
(författare)
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The structure of the second CysD domain of MUC2 and role in mucin organization by transglutaminase-based cross-linking
- 2024
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Ingår i: Cell Reports. - 2211-1247. ; 43
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Tidskriftsartikel (refereegranskat)abstract
- The MUC2 mucin protects the colonic epithelium by a two-layered mucus with an inner attached bacteria-free layer and an outer layer harboring commensal bacteria. CysD domains are 100 amino-acid-long sequences containing 10 cysteines that separate highly O-glycosylated proline, threonine, serine (PTS) regions in mucins. The structure of the second CysD, CysD2, of MUC2 is now solved by nuclear magnetic resonance. CysD2 shows a stable stalk region predicted to be partly covered by adjacent O-glycans attached to neighboring PTS sequences, whereas the CysD2 tip with three flexible loops is suggested to be well exposed. It shows transient dimer interactions at acidic pH, weakened at physiological pH. This transient interaction can be stabilized in vitro and in vivo by transglutaminase 3-catalyzed isopeptide bonds, preferring a specific glutamine residue on one flexible loop. This covalent dimer is modeled suggesting that CysD domains act as connecting hubs for covalent stabilization of mucins to form a protective mucus.
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| 7. |
- Trillo-Muyo, Sergio, 1983, et al.
(författare)
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Granule-stored MUC5B mucins are packed by the non-covalent formation of N-terminal head-to-head tetramers
- 2018
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Ingår i: Journal of Biological Chemistry. - : American Society for Biochemistry and Molecular Biology. - 0021-9258 .- 1083-351X. ; 293:15, s. 5746-5754
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Tidskriftsartikel (refereegranskat)abstract
- Most MUC5B mucin polymers in the upper airways of humans and pigs are produced by submucosal glands. MUC5B forms N-terminal covalent dimers that are further packed into larger assemblies because of low pH and high Ca2+ in the secretory granule of the mucin-producing cell. We purified the recombinant MUC5B N-terminal covalent dimer and used single-particle electron microscopy to study its structure under intracellular conditions. We found that, at intragranular pH, the dimeric MUC5B organized into head-to-head noncovalent tetramers where the von Willebrand D1-D2 domains hooked into each other. These N-terminal tetramers further formed long linear complexes from which, we suggest, the mucin domains and their C termini project radially outwards. Using conventional and video microscopy, we observed that, upon secretion into the submucosal gland ducts, a flow of bicarbonate-rich fluid passes the mucin-secreting cells. We suggest that this unfolds and pulls out the MUC5B assemblies into long linear threads. These further assemble into thicker mucin bundles in the glandular ducts before emerging at the gland duct opening. We conclude that the combination of intracellular packing of the MUC5B mucin and the submucosal gland morphology creates an efficient machine for producing linear mucin bundles.
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| 8. |
- Wahlgren, Weixiao Yuan, 1970, et al.
(författare)
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Structural mechanism of signal transduction in a phytochrome histidine kinase
- 2022
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Ingår i: Nature Communications. - : Springer Science and Business Media LLC. - 2041-1723. ; 13
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Tidskriftsartikel (refereegranskat)abstract
- Phytochrome proteins detect red/far-red light to guide the growth, motion, development and reproduction in plants, fungi, and bacteria. Bacterial phytochromes commonly function as an entrance signal in two-component sensory systems. Despite the availability of three-dimensional structures of phytochromes and other two-component proteins, the conformational changes, which lead to activation of the protein, are not understood. We reveal cryo electron microscopy structures of the complete phytochrome from Deinoccocus radiodurans in its resting and photoactivated states at 3.6 Å and 3.5 Å resolution, respectively. Upon photoactivation, the photosensory core module hardly changes its tertiary domain arrangement, but the connector helices between the photosensory and the histidine kinase modules open up like a zipper, causing asymmetry and disorder in the effector domains. The structures provide a framework for atom-scale understanding of signaling in phytochromes, visualize allosteric communication over several nanometers, and suggest that disorder in the dimeric arrangement of the effector domains is important for phosphatase activity in a two-component system. The results have implications for the development of optogenetic applications.
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