Increased understanding of the biochemistry and biosynthesis of MUC2 and other gel-forming mucins through the recombinant expression of their protein domains.
The gel-forming mucins are large and heavily O-glycosylated proteins which build up mucus gels. The recombinant production of full-length gel-forming mucins has not been possible to date. In order to study mucin biosynthesis and biochemistry, we and others have taken the alternative approach of constructing different recombinant proteins consisting of one or several domains of these large proteins and expressing them separately in different cell lines. Using this approach, we have determined that MUC2, the intestinal gel-forming mucin, dimerizes via its C-terminal cysteine-knot domain and also trimerizes via one of the N-terminal von Willebrand D domains. Both of these interactions are disulfide bond mediated. Via this assembly, a molecular network is built by which the mucus gel is formed. Here we discuss not only the functional understanding obtained from studies of the recombinant proteins, but also highlight the difficulties encountered when these proteins were produced recombinantly. We often found an accumulation of the proteins in the ER and consequently no secretion. This was especially apparent when the cysteine-rich domains of the N- and C-terminal parts of the mucins were expressed. Other proteins that we constructed were either not secreted or not expressed at all. Despite these problems, the knowledge of mucin biosynthesis and assembly has advanced considerably through the studies of these recombinant proteins.
Department of Medical Biochemistry, University of Gothenburg, Göteborg, Sweden. malin.backstrom@medkem.gu.se
Malin
M
Bäckström
Daniel
D
Ambort
Elisabeth
E
Thomsson
Malin E V
ME
Johansson
Gunnar C
GC
Hansson
D000818:Animals; D001671:Biochemistry; D016466:CHO Cells; D019556:COS Cells; D002460:Cell Line; D045744:Cell Line, Tumor; D002522:Chlorocebus aethiops; D003412:Cricetulus; D003545:Cysteine / Q000235:genetics / Q000378:metabolism; D019281:Dimerization; D005782:Gels / Q000378:metabolism; D006031:Glycosylation; D006801:Humans; D055262:Mucin-2 / Q000096:biosynthesis* / Q000235:genetics* / Q000378:metabolism; D017434:Protein Structure, Tertiary / Q000235:genetics*; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics* / Q000378:metabolism
D005782:Gels; D055262:Mucin-2; D011994:Recombinant Proteins; D003545:Cysteine