Clathrin light and heavy chain interface: α-helix binding superhelix loops via critical tryptophans

Chih Ying Chen, Michael L. Reese, Peter K. Hwang, Nobuyuki Ota, David Agard, Frances M. Brodsky

Research output: Contribution to journalArticle

38 Citations (Scopus)

Abstract

Clathrin light chain subunits (LCa and LCb) contribute to regulation of coated vesicle formation to sort proteins during receptor-mediated endocytosis and organelle biogenesis. LC binding to clathrin heavy chain (HC) was characterized by genetic and structural approaches. The core interactions were mapped to HC residues 1267-1522 (out of 1675) and LCb residues 90-157 (out of 228), using yeast two-hybrid assays. The C-termini of both subunits also displayed interactions extending beyond the core domains. Mutations to helix breakers within the LCb core disrupted HC association. Further suppressor mutagenesis uncovered compensatory mutations in HC (K1415E or K1326E) capable of rescuing the binding defects of LCb mutations W127R or W105R plus W138R, thereby pinpointing contacts between HC and LCb. Mutant HC K1415E also rescued loss of binding by LCa W130R, indicating that both LCs interact similarly with HC. Based on circular dichroism data, mapping and mutagenesis, LCa and LCb were represented as α-helices, aligned along the HC and, using molecular dynamics, a structural model of their interaction was generated with novel implications for LC control of clathrin assembly.

Original languageEnglish (US)
Pages (from-to)6072-6082
Number of pages11
JournalEMBO Journal
Volume21
Issue number22
DOIs
StatePublished - Nov 15 2002

Fingerprint

Clathrin Light Chains
Clathrin Heavy Chains
Mutagenesis
Tryptophan
Clathrin
Mutation
Yeast
Coated Vesicles
Molecular dynamics
Assays
Two-Hybrid System Techniques
Structural Models
Association reactions
Organelle Biogenesis
Molecular Dynamics Simulation
Circular Dichroism
Endocytosis
Defects
Proteins

Keywords

  • Cation-π interaction
  • Clathrin
  • Coat assembly
  • Molecular dynamics
  • Reverse two-hybrid screen

ASJC Scopus subject areas

  • Genetics
  • Cell Biology

Cite this

Clathrin light and heavy chain interface : α-helix binding superhelix loops via critical tryptophans. / Chen, Chih Ying; Reese, Michael L.; Hwang, Peter K.; Ota, Nobuyuki; Agard, David; Brodsky, Frances M.

In: EMBO Journal, Vol. 21, No. 22, 15.11.2002, p. 6072-6082.

Research output: Contribution to journalArticle

Chen, Chih Ying ; Reese, Michael L. ; Hwang, Peter K. ; Ota, Nobuyuki ; Agard, David ; Brodsky, Frances M. / Clathrin light and heavy chain interface : α-helix binding superhelix loops via critical tryptophans. In: EMBO Journal. 2002 ; Vol. 21, No. 22. pp. 6072-6082.
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