1. Biophysics Graduate Program, Department of Cell Biology, Harvard Medical School, 240 Longwood Avenue, Boston, Massachusetts 02115, USA
2. Department of Cell Biology, Harvard Medical School, 240 Longwood Avenue, Boston, Massachusetts 02115, USA
3. Howard Hughes Medical Institute and Department of Biochemistry, Rosenstiel Basic Medical Sciences Research Center, Brandeis University, 415 South Street, Waltham, Massachusetts 02454, USA
4. Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, and Howard Hughes Medical Institute, 250 Longwood Avenue, Boston, Massachusetts 02115, USA
5. Department of Cell Biology and the CBR Institute for Biomedical Research, Harvard Medical School, 200 Longwood Avenue, Boston, Massachusetts 02115, USA

Correspondence to: Tomas Kirchhausen⁵ Email: Kirchhausen@crystal.harvard.edu
Coordinates have been deposited in the Protein Data Bank under accession number 1XI5.

Abstract

Clathrin-coated pits invaginate from specific membrane compartments and pinch off as coated vesicles. These vesicles then uncoat rapidly once released. The Hsc70 molecular chaperone effects the uncoating reaction, and is guided to appropriate locations on clathrin lattices by the J-domain-containing co-chaperone molecule auxilin^{1, 2, 3, 4}. This raises the question of how a local event such as ATP hydrolysis by Hsc70 can catalyse a global disassembly. Here, we have used electron cryomicroscopy to determine 12-Å-resolution structures of in-vitro-assembled clathrin coats in association with a carboxy-terminal fragment of auxilin that contains both the clathrin-binding region and the J domain. We have located the auxilin fragment by computing differences between these structures and those lacking auxilin (described in an accompanying paper⁵). Auxilin binds within the clathrin lattice near contacts between an inward-projecting C-terminal helical tripod and the crossing of two 'ankle' segments; it also contacts the terminal domain of yet another clathrin 'leg'. It therefore recruits Hsc70 to the neighbourhood of a set of critical interactions. Auxilin binding produces a local change in heavy-chain contacts, creating a detectable global distortion of the clathrin coat. We propose a mechanism by which local destabilization of the lattice promotes general uncoating.

1. Biophysics Graduate Program, Department of Cell Biology, Harvard Medical School, 240 Longwood Avenue, Boston, Massachusetts 02115, USA
2. Department of Cell Biology, Harvard Medical School, 240 Longwood Avenue, Boston, Massachusetts 02115, USA
3. Howard Hughes Medical Institute and Department of Biochemistry, Rosenstiel Basic Medical Sciences Research Center, Brandeis University, 415 South Street, Waltham, Massachusetts 02454, USA
4. Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, and Howard Hughes Medical Institute, 250 Longwood Avenue, Boston, Massachusetts 02115, USA
5. Department of Cell Biology and the CBR Institute for Biomedical Research, Harvard Medical School, 200 Longwood Avenue, Boston, Massachusetts 02115, USA

Correspondence to: Tomas Kirchhausen⁵ Email: Kirchhausen@crystal.harvard.edu
Coordinates have been deposited in the Protein Data Bank under accession number 1XI5.

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