PDBsum entry 3mxw

Signaling protein

PDB id: 3mxw

Contents

Protein chains

153 a.a. *

212 a.a. *

220 a.a. *

Ligands

SO4 ×3

Metals

_ZN

_CA ×2

Waters ×423

* Residue conservation analysis

PDB id:

3mxw

Name:

Signaling protein

Title:

Crystal structure sonic hedgehog bound to the 5e1 fab fragment

Structure:

Sonic hedgehog protein. Chain: a. Synonym: shh, hhg-1, sonic hedgehog protein n-product, sonic hedgehog protein c-product. Engineered: yes. 5e1 light chain. Chain: l. Engineered: yes. 5e1 heavy chain.

Source:

Homo sapiens. Human. Organism_taxid: 9606. Gene: shh. Expressed in: escherichia coli. Expression_system_taxid: 562. Promoter. Expression_system_taxid: 562

Resolution:

1.83Å R-factor: 0.181 R-free: 0.218

Authors:

S.G.Hymowitz,H.R.Maun

Key ref:

H.R.Maun et al. (2010). Hedgehog pathway antagonist 5E1 binds hedgehog at the pseudo-active site. J Biol Chem, 285, 26570-26580. PubMed id: 20504762

Date:

07-May-10 Release date: 26-May-10

Protein chain

P61964  (WDR5_HUMAN) -  WD repeat-containing protein 5 from Homo sapiens

Seq: 334 a.a.

Struc: 153 a.a.*

Protein chain

No UniProt id for this chain

Struc: 212 a.a.

Protein chain

No UniProt id for this chain

Struc: 220 a.a.

* PDB and UniProt seqs differ at 142 residue positions (black crosses)

Enzyme reactions

• Enzyme class: Chain A: E.C.3.1.-.-

J Biol Chem 285:26570-26580 (2010)

PubMed id: 20504762

Hedgehog pathway antagonist 5E1 binds hedgehog at the pseudo-active site.

H.R.Maun, X.Wen, A.Lingel, F.J.de Sauvage, R.A.Lazarus, S.J.Scales, S.G.Hymowitz.

ABSTRACT

Proper hedgehog (Hh) signaling is crucial for embryogenesis and tissue regeneration. Dysregulation of this pathway is associated with several types of cancer. The monoclonal antibody 5E1 is a Hh pathway inhibitor that has been extensively used to elucidate vertebrate Hh biology due to its ability to block binding of the three mammalian Hh homologs to the receptor, Patched1 (Ptc1). Here, we engineered a murine:human chimeric 5E1 (ch5E1) with similar Hh-binding properties to the original murine antibody. Using biochemical, biophysical and X-ray crystallographic studies, we show that, like the regulatory receptors Cdon and Hedgehog-interacting protein (Hhip), ch5E1 binding to Sonic hedgehog (Shh) is enhanced by calcium ions. In the presence of calcium and zinc ions, the ch5E1 binding affinity increases 10 to 20-fold to tighter than 1 nM primarily due to decrease in the dissociation rate. The co-crystal structure of Shh bound to the Fab fragment of ch5E1 reveals that 5E1 binds at the pseudo-active site groove of Shh with an epitope that largely overlaps with the binding site of its natural receptor antagonist Hhip. Unlike Hhip, the side chains of 5E1 do not directly coordinate the Zn2+ cation in the pseudo-active site, despite the modest zinc-dependent increase in 5E1 affinity for Shh. Furthermore, to our knowledge, the ch5E1 Fab-Shh complex represents the first structure of an inhibitor antibody bound to a metalloprotease fold.