Unpacking a gel-forming mucin: a view of MUC5B organization after granular release

Abstract

Kesimer M, Makhov AM, Griffith JD, Verdugo P, Sheehan JK. Unpacking a gel-forming mucin: a view of MUC5B organization after granular release. Am J Physiol Lung Cell Mol Physiol 298: L15-L22, 2010. First published September 25, 2009 doi:10.1152/ajplung.00194.2009.-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 ϳ150 ϫ 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 NH2-and COOH-terminal globular protein domains within the granule and also explains how the mucin can expand so rapidly upon its release. mucin granule; secretion; mucus MUCINS ARE LARGE COMPLEX GLYCOPROTEIN macromolecules that provide the basis of gels protecting the surface epithelia in many multicellular organisms. In humans, these macromolecules come in two major kinds, the epithelial or transmembrane mucins characterized by COOH-terminal domains that attach to cell surface membranes (5) and those termed the gel-forming mucins that are secreted as highly assembled polymeric structures (14). There are four such molecules, MUC2, MUC5AC, MUC5B, and MUC6, and they form the largest glycoproteins in the body (10). The functions of these molecules are understood in general terms, i.e., they form the basic infrastructure of the epithelial protective gels, but the mechanisms by which they are made and organized within cells and thereafter released are not understood.The basis of the polymeric assembly of these large mucin glycoproteins is thought to mimic that of another related glycoprotein calle...

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