Dynamic Interactions between Clathrin and Locally Structured Elements in a Disordered Protein Mediate Clathrin Lattice Assembly

Yue Zhuo, Udayar Ilangovan, Virgil Schirf, Borries Demeler, Rui Sousa, Andrew P. Hinck, Eileen M. Lafer

Research output: Contribution to journalArticle

32 Scopus citations

Abstract

Assembly of clathrin lattices is mediated by assembly/adaptor proteins that contain domains that bind lipids or membrane-bound cargo proteins and clathrin binding domains (CBDs) that recruit clathrin. Here, we characterize the interaction between clathrin and a large fragment of the CBD of the clathrin assembly protein AP180. Mutational, NMR chemical shift, and analytical ultracentrifugation analyses allowed us to precisely define two clathrin binding sites within this fragment, each of which is found to bind weakly to the N-terminal domain of the clathrin heavy chain (TD). The locations of the two clathrin binding sites are consistent with predictions from sequence alignments of previously identified clathrin binding elements and, by extension, indicate that the complete AP180 CBD contains ̃ 12 degenerate repeats, each containing a single clathrin binding site. Sequence and circular dichroism analyses have indicated that the AP180 CBD is predominantly unstructured and our NMR analyses confirm that this is largely the case for the AP180 fragment characterized here. Unexpectedly, unlike the many proteins that undergo binding-coupled folding upon interaction with their binding partners, the AP180 fragment is similarly unstructured in its bound and free states. Instead, we find that this fragment exhibits localized β-turn-like structures at the two clathrin binding sites both when free and when bound to clathrin. These observations are incorporated into a model in which weak binding by multiple, pre-structured clathrin binding elements regularly dispersed throughout a largely unstructured CBD allows efficient recruitment of clathrin to endocytic sites and dynamic assembly of the clathrin lattice.

Original languageEnglish (US)
Pages (from-to)274-290
Number of pages17
JournalJournal of Molecular Biology
Volume404
Issue number2
DOIs
StatePublished - Nov 26 2010

Keywords

  • AP180
  • Clathrin
  • Endocytosis
  • Intrinsically disordered protein
  • Intrinsically unstructured protein

ASJC Scopus subject areas

  • Structural Biology
  • Molecular Biology

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