Assembly of the respiratory Mucin MUC5B a new model for a gel-forming Mucin

Research output: Contribution to journalArticle

  • External authors:
  • Nikos Kouvatsos
  • Bertrand D. Raynal
  • Marj Howard
  • Richard F. Collins
  • Jean Luc Desseyn
  • C. William Davis

Abstract

Mucins are essential components in mucus gels thatform protective barriers at all epithelial surfaces, but much remains unknown about their assembly, intragranular organization, and post-secretion unfurling to form mucus. MUC5B is a major polymeric mucin expressed by respiratory epithelia, and we investigated the molecular mechanisms involved during its assembly. Studies of intact polymeric MUC5B revealed a single high affinity calcium-binding site, distinct from multiple low affinity sites on each MUC5B monomer. Self-diffusion studies with intact MUC5B showed that calcium binding at the protein site catalyzed reversible cross-links between MUC5B chains to form networks. The site of cross-linking was identified in the MUC5B D3-domain as it was specifically blocked by D3 peptide antibodies. Biophysical analysis and single particle EM of recombinant MUC5B N terminus (D1D2D'D3; NT5B) and subdomains (D1, D1-D2, D2-D'-D3, and D3) generated structural models of monomers and disulfide-linked dimers and suggested that MUC5B multimerizes by disulfide linkage between D3-domains to form linear polymer chains. Moreover, these analyses revealed reversible homotypic interactions of NT5B at low pH and in high calcium, between disulfide-linked NT5B dimers, but not monomers. These results enable a model of MUC5B to be derived, which predicts mechanisms of mucin intracellular assembly and storage, which may be common to the other major gel-forming polymeric mucins. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc. Published in the U.S.A.

Bibliographical metadata

Original languageEnglish
Pages (from-to)16409-16420
Number of pages11
JournalJournal of Biological Chemistry
Volume289
Issue number23
DOIs
Publication statusPublished - 6 Jun 2014

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