PDBsum entry 2qc9

Transcription

PDB id: 2qc9

Contents

Protein chains

193 a.a. *

Waters ×442

* Residue conservation analysis

PDB id:

2qc9

Name:

Transcription

Title:

Mouse notch 1 ankyrin repeat intracellular domain

Structure:

Notch 1 protein. Chain: a, b. Fragment: ankyrin repeat domain. Engineered: yes

Source:

Mus musculus. House mouse. Organism_taxid: 10090. Gene: notch1. Expressed in: escherichia coli bl21(de3). Expression_system_taxid: 469008.

Resolution:

2.35Å R-factor: 0.173 R-free: 0.228

Authors:

M.A.Mcdonough,C.J.Schofield

Key ref:

M.L.Coleman et al. (2007). Asparaginyl hydroxylation of the Notch ankyrin repeat domain by factor inhibiting hypoxia-inducible factor. J Biol Chem, 282, 24027-24038. PubMed id: 17573339 DOI: 10.1074/jbc.M704102200

Date:

19-Jun-07 Release date: 04-Mar-08

Protein chains

Q01705  (NOTC1_MOUSE) -  Neurogenic locus notch homolog protein 1 from Mus musculus

Seq: 2531 a.a.

Struc: 193 a.a.*

* PDB and UniProt seqs differ at 2 residue positions (black crosses)

DOI no: 10.1074/jbc.M704102200

J Biol Chem 282:24027-24038 (2007)

PubMed id: 17573339

Asparaginyl hydroxylation of the Notch ankyrin repeat domain by factor inhibiting hypoxia-inducible factor.

M.L.Coleman, M.A.McDonough, K.S.Hewitson, C.Coles, J.Mecinovic, M.Edelmann, K.M.Cook, M.E.Cockman, D.E.Lancaster, B.M.Kessler, N.J.Oldham, P.J.Ratcliffe, C.J.Schofield.

ABSTRACT

The stability and activity of hypoxia-inducible factor (HIF) are regulated by the post-translational hydroxylation of specific prolyl and asparaginyl residues. We show that the HIF asparaginyl hydroxylase, factor inhibiting HIF (FIH), also catalyzes hydroxylation of highly conserved asparaginyl residues within ankyrin repeat (AR) domains (ARDs) of endogenous Notch receptors. AR hydroxylation decreases the extent of ARD binding to FIH while not affecting signaling through the canonical Notch pathway. ARD proteins were found to efficiently compete with HIF for FIH-dependent hydroxylation. Crystallographic analyses of the hydroxylated Notch ARD (2.35A) and of Notch peptides bound to FIH (2.4-2.6A) reveal the stereochemistry of hydroxylation on the AR and imply that significant conformational changes are required in the ARD fold in order to enable hydroxylation at the FIH active site. We propose that ARD proteins function as natural inhibitors of FIH and that the hydroxylation status of these proteins provides another oxygen-dependent interface that modulates HIF signaling.

Selected figure(s)

Figure 5.
Structure of Asn-hydroxylated N1 ARD.A, stereoview ribbon representation from the 2.35 Å resolution crystal structure of N1 ARD (OH); six ARs (numbered 2–7) are observed in the N1 ARD (OH) structure. The hydroxylated Asn-1945 (AR 2) and unhydroxylated Asn-2012 (AR 4) are yellow ball-and-stick representations. B, stereoview of electron density around Asn-1945 showing the pro-S configuration of the hydroxylated Asn-1945 β-carbon. 2F[o] - F[c] map (cyan mesh) contoured to 1σ and an F[o] - F[c] OMIT map (dark blue mesh) contoured to 5σ (produced by omitting the hydroxyl group from the calculation). This region of the β-hairpin loop has a type I β-turn (Asp-1943, Ala-1944, Asn-1945, and Ile-1946 at the i, i + 1, i + 2, and i + 3 positions, respectively). Asn-1945 makes three hydrogen bonds (dashed black lines); the nitrogen and oxygen of the side chain amide form hydrogen bonds to the backbone carbonyl oxygen of Ala-1975 and NH of Asp-1977, and the β-hydroxy group hydrogen bonds with an Asp-1943 carboxylate oxygen. C, ribbon representation of the Asn-1945 hydroxylated mN1-(1898–2105) (blue) and the MAML-1 (green)·human N1-(1873–2127) (purple)·CSL (yellow/beige/orange)·DNA (blue) complex (Protein Data Bank code 2F8X) structures (24) superimposed.

Figure 6.
Notch-FIH interactions.A, the FIH homodimer (molecule A (pink) and molecule B (green)) in complex with N1-(1930–1949) (yellow ball-and-stick representation bound to A, blue bound to B) with the double-stranded β-helix core (blue in A and raspberry in B) and Fe(II) (orange sphere). B, stereoview ribbon representation of the FIH·Fe(II)·2OG·N1-(1930–1949) dimer structure complexed with the N1 peptide to 2.4 Å resolution in yellow ball-and-stick representation and the σ[A] weighted composite OMIT mF[o] - DF[c] difference electron density contoured to 3σ (blue mesh) created by omitting the Notch1 atoms from the calculation. Electron density was apparent for residues 1936–1945 of the N1-(1930–1949) peptide (close up view; supplemental Fig. S3). C, the FIH active site (FIH (salmon with residues in white ball-and-stick representation), 2OG (green), N1-(1930–1949) (yellow), and Fe(II) (orange sphere)). The pro-S C–H bond (gray) modeled at the Cβ position of N1 Asn-1945 is positioned for oxidation. D, ball-and-stick representation of the superimposed peptides when bound to FIH. yellow, N1-(1930–1949); cyan, N1-(1997–2016); white, HIF1αCAD. E, close up of superimposed structures: FIH·N1-(1930–1949) (yellow), FIH·N1-(1997–2016) (cyan), and FIH·HIF1αCAD (white) showing the binding of the conserved leucine (N1 Leu-1937/2004 and HIF1αCAD Leu-795) in a hydrophobic pocket (yellow-green surface) formed by residues from molecules A (dark green) and B(orange) of the FIH dimer. F, superimposition of the target Asn of N1-(1930–1949) when bound to FIH and the equivalent Asn in the N1 ARD structure emphasizes the conformational changes between them.

The above figures are reprinted by permission from the ASBMB: J Biol Chem (2007, 282, 24027-24038) copyright 2007.

Figures were selected by an automated process.