Crystal of hypoxia inducible factor 1 alpha prolyl hydroxylase
View Patent ↗The crystal structure of ligand-bound EGLN1 catalytic domain of prolyl hydroxylase is disclosed. These coordinates are useful in computer aided drug design for identifying compounds that regulate EGLN1 prolyl hydroxylase and thereby regulate HIF-regulated disorders.
1. A method for identifying a compound that binds to Hypoxia Inducible Factor-1 alpha prolyl hydroxylase (EGLN1), comprising:
a) providing a crystal of an EGLN1 catalytic domain polypeptide comprising the amino acid sequence of SEQ ID NO: 3 in complex with [(1-chloro-4-hydroxy-isoquinoline-3-carbonyl)amino]acetic acid, wherein the crystal has unit cell dimensions of a=111±1 Å, b=111±1 Å, and c=40±1 Å, α=90°, β=90°, γ=120° in the hexagonal space group P6 3 ;
b) exposing the crystal to a compound in a medium to form a crystal/compound complex;
c) irradiating the crystal/compound complex with X-rays to generate a diffraction pattern;
d) capturing the pattern to a recording device to generate diffraction data;
e) processing the data to solve the structure of the crystal/compound complex; and
f) determining the location and binding geometry of the compound within the structure of the complex;
thereby identifying a compound that binds to EGLN1.
2. The method according to claim 1 , further comprising:
g) selecting a candidate compound that has been shown to bind to EGLN1; and
h) determining if the candidate compound binds EGLN1 in an in vitro, in vivo, or ex vivo assay.
3. The method according to claim 2 , wherein determining if the compound binds EGLN1 in step (h) is performed using an enzymatic assay.
4. The method according to claim 2 , wherein determining if the compound binds EGLN1 in step (h) is performed using a cell-based assay.
5. A method for identifying a compound that binds to Hypoxia Inducible Factor-1 alpha prolyl hydroxylase (EGLN1), comprising:
a) providing a crystal of an EGLN1 catalytic domain polypeptide comprising the amino acid sequence of SEQ ID NO: 3 in complex with [(1-chloro-4-hydroxy-isoquinoline-3-carbonyl)amino]acetic acid, wherein the crystal has unit cell dimensions of a=111±1 Å, b=111±1 Å, and c=40±1 Å, α=90°, β=90°, γ=120° in the hexagonal space group P6 3 ;
b) exposing the crystal to a candidate compound in a medium to form a crystal/compound complex;
c) irradiating the crystal/compound complex with X-rays to generate a diffraction pattern;
d) capturing the pattern to a recording device to generate diffraction data;
e) processing the data to solve the structure of the crystal/compound complex;
f) determining the location and binding geometry of the compound within the structure of the complex;
g) selecting a candidate compound that has been shown to bind to EGLN1; and
h) determining whether the candidate compound binds to EGLN1 in an in vitro or in vivo assay.
6. The method according to claim 5 , wherein an enzyme-based assay is used in step (h) for determining whether the candidate compound binds to EGLN1.
7. The method according to claim 5 , wherein a cell-based assay is used in step (h) for determining whether the candidate compound binds to EGLN1.