Unpacking a gel-forming mucin: a view of MUC5B organization after granular release.
Gel-forming mucins are the largest complex glycoprotein macromolecules in the body. They form the matrix of gels protecting all the surface epithelia and are secreted as disulfide-bonded polymeric structures. The mechanisms by which they are formed and organized within cells and thereafter released to form mucus gels are not understood. In particular, the initial rate of expansion of the mucins after release from their secretory granules is very rapid (seconds), but no clear mechanism for how it is achieved has emerged. Our major interest is in lung mucins, but most particularly in MUC5B, which is the major gel-forming mucin in mucus, and which provides its major protective matrix. In this study, using OptiPrep density gradient ultracentrifugation, we have isolated a small amount of a stable form of the recently secreted and expanding MUC5B mucin, which accounts for less than 2% of the total mucin present. It has an average mass of approximately 150 x 10(6) Da and size Rg of 150 nm in radius of gyration. In transmission electron microscopy, this compact mucin has maintained a circular structure that is characterized by flexible chains connected around protein-rich nodes as determined by their ability to bind colloidal gold. The appearance indicates that the assembled mucins in a single granular form are organized around a number of nodes, each attached to four to eight subunits. The organization of the mucins in this manner is consistent with efficient packing of a number of large heavily glycosylated monomers while still permitting their rapid unfolding and hydration. For the first time, this provides some insight into how the carbohydrate regions might be organized around the NH(2)- and COOH-terminal globular protein domains within the granule and also explains how the mucin can expand so rapidly upon its release.
Kesimer
Mehmet
M
1Department of Biochemistry and Biophysics and Cystic Fibrosis/Pulmonary Research and Treatment Center, Chapel Hill, NC, USA. kesimer@med.unc.edu
Makhov
Alexander M
AM
Griffith
Jack D
JD
Verdugo
Pedro
P
Sheehan
John K
JK
D005782:Gels / Q000378:metabolism*; D006801:Humans; D008297:Male; D008958:Models, Molecular; D008970:Molecular Weight; D055257:Mucin-5B / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism* / Q000648:ultrastructure; D017434:Protein Structure, Tertiary; D022142:Secretory Vesicles / Q000378:metabolism*
D005782:Gels; D055257:Mucin-5B