PDBsum entry 1h0a

Endocytosis

PDB id: 1h0a

Contents

Protein chain

158 a.a. *

Ligands

DIO ×5

I3P

Waters ×204

* Residue conservation analysis

PDB id:

1h0a

Name:

Endocytosis

Title:

Epsin enth bound to ins(1,4,5)p3

Structure:

Epsin. Chain: a. Fragment: enth domain, residues 1-158. Engineered: yes

Source:

Rattus norvegicus. Rat. Organism_taxid: 10116. Expressed in: escherichia coli. Expression_system_taxid: 511693. Expression_system_variant: codon plus.

Resolution:

1.70Å R-factor: 0.189 R-free: 0.228

Authors:

M.G.J.Ford,H.T.Mcmahon,P.R.Evans

Key ref:

M.G.Ford et al. (2002). Curvature of clathrin-coated pits driven by epsin. Nature, 419, 361-366. PubMed id: 12353027 DOI: 10.1038/nature01020

Date:

12-Jun-02 Release date: 03-Oct-02

Protein chain

O88339  (EPN1_RAT) -  Epsin-1 from Rattus norvegicus

Seq: 575 a.a.

Struc: 158 a.a.

DOI no: 10.1038/nature01020

Nature 419:361-366 (2002)

PubMed id: 12353027

Curvature of clathrin-coated pits driven by epsin.

M.G.Ford, I.G.Mills, B.J.Peter, Y.Vallis, G.J.Praefcke, P.R.Evans, H.T.McMahon.

ABSTRACT

Clathrin-mediated endocytosis involves cargo selection and membrane budding into vesicles with the aid of a protein coat. Formation of invaginated pits on the plasma membrane and subsequent budding of vesicles is an energetically demanding process that involves the cooperation of clathrin with many different proteins. Here we investigate the role of the brain-enriched protein epsin 1 in this process. Epsin is targeted to areas of endocytosis by binding the membrane lipid phosphatidylinositol-4,5-bisphosphate (PtdIns(4,5)P(2)). We show here that epsin 1 directly modifies membrane curvature on binding to PtdIns(4,5)P(2) in conjunction with clathrin polymerization. We have discovered that formation of an amphipathic alpha-helix in epsin is coupled to PtdIns(4,5)P(2) binding. Mutation of residues on the hydrophobic region of this helix abolishes the ability to curve membranes. We propose that this helix is inserted into one leaflet of the lipid bilayer, inducing curvature. On lipid monolayers epsin alone is sufficient to facilitate the formation of clathrin-coated invaginations.

Selected figure(s)

Figure 1.
Figure 1: Epsin 1 ENTH domain tubulates liposomes. a, Modular arrangement of endocytic proteins. The epsin family can be recognized by the presence of an N-terminal lipid-binding ENTH domain and a clathrin/adaptor-binding domain. The ENTH domain is homologous over the first 150 residues to the ANTH domains of AP180, CALM and HIP1, although the lipid-binding residues are very different (see Fig. 2d). The ENTH domain is followed by ubiquitin-interacting motifs. The adaptor-binding motifs in epsin 1 (eight DPW motifs) and DPF-like motifs in other endocytic proteins bind to the appendage domains of the AP2 complex25. NPF motifs at the C terminus of epsin 1 interact with the Eps15 homology (EH) domain of Eps15 (refs 7 -9). The scale bar is in amino acid residues. b, Electron microscopy of liposomes in the presence of the domains indicated. dAmphN, Drosophila amphiphysin N-terminus26. The outer diameter of tubules formed with epsin ENTH domain was 15 1 nm, that of tubules formed with full-length epsin was 19 3 nm, and that in the presence of dAmphN was 46 2 nm.

Figure 2.
Figure 2: Structure of epsin ENTH bound to Ins(1,4,5)P[3]. a, Ribbon diagrams of epsin ENTH bound to Ins(1,4,5)P[3] (Protein DataBank (PDB) accession number 1H0A), and for comparison the previous structures of epsin ENTH solved in the absence of Ins(1,4,5)P[3] (PDB 1EDU14) and CALM ANTH bound to diC[8]PtdIns(4,5)P[2] (PDB 1HFA^13). The structures are coloured red to blue from N- to C-termini, with corresponding helices having the same colour. Surface electrostatic potentials (red - 10 kT e^-1; blue + 10 kT e^-1) of each structure are shown below. Ins(1,4,5)P[3] binds to CALM on a positively charged surface not present in epsin. b, Schematic diagram of the interactions responsible for binding the Ins(1,4,5)P[3] molecule. c, Close-up view of the Ins(1,4,5)P[3] binding site, showing the residues responsible for interaction with the ligand. The hydrophobic residues L6, M10 and I13 exposed on formation of helix 0 are also marked. The structure shows that lipid binding and an interaction of helix 0 with the bilayer could happen simultaneously. The electron density for the ligand is shown, contoured at 0.168 e �-3. d, Sequence alignments comparing the lipid-binding residues of all the epsin family members with corresponding residues from the ANTH domains of AP180, CALM and LAP (the Drosophila AP180 homologue). Critical residues for Ins(1,4,5)P[3] binding to epsin 1 are coloured in blue and are conserved in epsins 1, 2 and 3 and in Drosophila epsin (liquid facets). The lipid-binding residues are not well conserved in epsinR/Drosophila epsin-like (see residues coloured green), suggesting a different lipid specificity of this epsin. The epsin Ins(1,4,5)P[3]-binding residues are not conserved in AP180 and CALM, where a different set of residues have been identified as being involved in Ins(1,4,5)P[3] binding, coloured in purple^13. h, human; d, Drosophila; r, rat.

The above figures are reprinted by permission from Macmillan Publishers Ltd: Nature (2002, 419, 361-366) copyright 2002.

Figures were selected by the author.

Author's comment

The ENTH domain binds to PtdIns(4,5)P2. On binding to the lipid, the otherwise unstructured N-terminal residues fold into a helix (termed helix 0 to keep with the naming convention of the previously solved ENTH domain - pdb id 1EDU). Helix 0 is amphipathic and the hydrophobic residues on the outer surface of helix 0 can insert into PtdIns(4,5)P2-containing membranes and function like a wedge to generate curvature. In in vitro experiments using liposomes, the result is extensive tubulation and in the presence of an epsin binding partner, clathrin, it generates clathrin-coated pits. Though the ENTH domain shares some sequence conservation with the ANTH domain (pdb id 1HFA), and both bind to PtdIns(4,5)P2, yet the modes of binding, and the functional consequences thereof, are very different.
Harvey McMahon Marijn Ford