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Edited by Linda L. Randall, University of Missouri-Columbia, Columbia, MO, and approved September 1, 2017 (received for review February 1, 2017)
Significance
How protein assemblies with complex topologies are formed is an important question in structural biology. An intriguing example is IgM, a complex of over 1,200 kDa consisting of six antibody subunits (or five in the presence of the J-chain protein). These are arranged in a ring-like structure connected by disulfide bonds. Here, we show that in vitro and in cell culture, a short peptide extension of the IgM heavy chain is sufficient to steer the formation of the hexameric complex. The formation of a disulfide bond triggers conformational changes in the peptide extensions, which involve specific hydrophobic residues. Our study reveals the redox-controlled assembly of a large protein complex via structural rearrangements in a peptide as a design principle.
Abstract
Professional secretory cells can produce large amounts of high-quality complex molecules, including IgM antibodies. Owing to their multivalency, polymeric IgM antibodies provide an efficient first-line of defense against pathogens. To decipher the mechanisms of IgM assembly, we investigated its biosynthesis in living cells and faithfully reconstituted the underlying processes in vitro. We find that a conserved peptide extension at the C-terminal end of the IgM heavy (Ig-μ) chains, termed the tailpiece, is necessary and sufficient to establish the correct geometry. Alanine scanning revealed that hydrophobic amino acids in the first half of the tailpiece contain essential information for generating the correct topology. Assembly is triggered by the formation of a disulfide bond linking two tailpieces. This induces conformational changes in the tailpiece and the adjacent domain, which drive further polymerization. Thus, the biogenesis of large and topologically challenging IgM complexes is dictated by a local conformational switch in a peptide extension.
Footnotes
↵1Present address: Interaction Analytics, Protein Sciences & CMC Department, Morphosys AG, 82152 Planegg, Germany.
- ↵2To whom correspondence may be addressed. Email: sitia.roberto{at}hsr.it or johannes.buchner{at}tum.de.
Author contributions: D.P., R.M., C.F.W.B., R.S., and J.B. designed research; D.P., B.W., C.G., T.A., R.M., C.F., M.F., C.J., and M.F.M. performed research; D.P., B.W., and C.G. analyzed data; and D.P., B.W., T.A., R.S., and J.B. wrote the paper.
The authors declare no conflict of interest.
This article is a PNAS Direct Submission.
This article contains supporting information online at www.pnas.org/lookup/suppl/doi:10.1073/pnas.1701797114/-/DCSupplemental.
Peptide extension dictates IgM assembly
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