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The C-terminal dimerization domain of the respiratory mucin MUC5B functions in mucin stability and intracellular packaging before secretion

DOI: 10.1074/jbc.RA119.010771 DOI Help

Authors: Caroline Ridley (Th University of Manchester) , Michael P. Lockhart-cairns (The University of Manchester) , Richard F. Collins (The University of Manchester) , Thomas A. Jowitt (The University of Manchester) , Durai B. Subramani (University of North Carolina School of Medicine) , Mehmet Kesimer (University of North Carolina School of Medicine) , Clair Baldock (The University of Manchester) , David J. Thornton (The University of Manchester)
Co-authored by industrial partner: No

Type: Journal Paper
Journal: Journal Of Biological Chemistry , VOL 294 , PAGES 17105 - 17116

State: Published (Approved)
Published: November 2019
Diamond Proposal Number(s): 16619 , 17773

Open Access Open Access

Abstract: Mucin 5B (MUC5B) has an essential role in mucociliary clearance that protects the pulmonary airways. Accordingly, knowledge of MUC5B structure and its interactions with itself and other proteins is critical to better understand airway mucus biology and improve the management of lung diseases such as asthma, cystic fibrosis, and chronic obstructive pulmonary disease (COPD). The role of an N-terminal multimerization domain in the supramolecular organization of MUC5B has been previously described, but less is known about its C-terminal dimerization domain. Here, using cryogenic electron microscopy (cryo-EM) and small-angle X-ray scattering (SAXS) analyses of recombinant disulfide-linked dimeric MUC5B dimerization domain we identified an asymmetric, elongated twisted structure, with a double globular base. We found that the dimerization domain is more resistant to disruption than the multimerization domain suggesting the twisted structure of the dimerization domain confers additional stability to MUC5B polymers. Size-exclusion chromatography-multiangle light scattering (SEC-MALS), SPR-based biophysical analyses and microscale thermophoresis of the dimerization domain disclosed no further assembly, but did reveal reversible, calcium-dependent interactions between the dimerization and multimerization domains that were most active at acidic pH, suggesting that these domains have a role in MUC5B intragranular organization. In summary, our results suggest a role for the C-terminal dimerization domain of MUC5B in compaction of mucin chains during granular packaging via interactions with the N-terminal multimerization domain. Our findings further suggest that the less stable multimerization domain provides a potential target for mucin depolymerization to remove mucus plugs in COPD and other lung pathologies.

Journal Keywords: mucin; mucus; von Willebrand factor; lung; cryo-electron microscopy; small-angle X-ray scattering (SAXS); Mucus obstruction

Subject Areas: Biology and Bio-materials

Diamond Offline Facilities: Electron Bio-Imaging Centre (eBIC)
Instruments: B21-High Throughput SAXS , Krios I-Titan Krios I at Diamond

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