Cellular oxygen sensing: Crystal structure of hypoxia-inducible factor prolyl hydroxylase (PHD2)

doi:10.1073/pnas.0601283103

Cellular oxygen sensing: Crystal structure of hypoxia-inducible factor prolyl hydroxylase (PHD2)

*Oxford Centre for Molecular Sciences and Department of Chemistry, University of Oxford, Mansfield Road, Oxford OX1 3TA, United Kingdom;
^§ReOx Ltd., Magdalen Center, Oxford Science Park, Oxford OX4 4GA, United Kingdom; and
^‡Amgen, Inc., One Amgen Center Drive, Thousand Oaks, CA 91320-1789

Edited by Peter B. Dervan, California Institute of Technology, Pasadena, CA, and approved May 17, 2006
↵ ^†M.A.M. and R.S.S. contributed equally to this work. (received for review February 15, 2006)

Abstract

Cellular and physiological responses to changes in dioxygen levels in metazoans are mediated via the posttranslational oxidation of hypoxia-inducible transcription factor (HIF). Hydroxylation of conserved prolyl residues in the HIF-α subunit, catalyzed by HIF prolyl-hydroxylases (PHDs), signals for its proteasomal degradation. The requirement of the PHDs for dioxygen links changes in dioxygen levels with the transcriptional regulation of the gene array that enables the cellular response to chronic hypoxia; the PHDs thus act as an oxygen-sensing component of the HIF system, and their inhibition mimics the hypoxic response. We describe crystal structures of the catalytic domain of human PHD2, an important prolyl-4-hydroxylase in the human hypoxic response in normal cells, in complex with Fe(II) and an inhibitor to 1.7 Å resolution. PHD2 crystallizes as a homotrimer and contains a double-stranded β-helix core fold common to the Fe(II) and 2-oxoglutarate-dependant dioxygenase family, the residues of which are well conserved in the three human PHD enzymes (PHD 1–3). The structure provides insights into the hypoxic response, helps to rationalize a clinically observed mutation leading to familial erythrocytosis, and will aid in the design of PHD selective inhibitors for the treatment of anemia and ischemic disease.

Footnotes

^¶To whom correspondence may be addressed. E-mail: rsyed{at}amgen.com or christopher.schofield{at}chem.ox.ac.uk
Author contributions: M.A.M., K.S.H., R.S.S., and C.J.S. designed research; M.A.M., V.L., E.F., R.C., C.M., B.M.R.L., J.Z., N.J.O., I.J.C., J.L., L.A.M., R.J.M.K., K.S.H., E.Y., and R.S.S. performed research; M.A.M., V.L., J.Z., L.A.M., R.J.M.K., and K.S.H. contributed new reagents/analytic tools; M.A.M., E.F., B.M.R.L., S.J., R.S.S., and C.J.S. analyzed data; and M.A.M., R.S.S., and C.J.S. wrote the paper.
Conflict of interest statement: C.J.S. is a cofounder and K.S.H. and L.A.M. are employees of ReOx, Ltd., Oxford, and R.S.S., V.L., J.Z., J.L., R.J.M.K., E.Y., and S.J. are employees of Amgen, Inc. Both ReOX and Amgen are developing therapeutic inhibitors of the HIF hydroxylases.
This paper was submitted directly (Track II) to the PNAS office.
Data deposition: The atomic coordinates have been deposited in the Protein Data Bank, www.pdb.org (PDB ID codes 2G19 and 2G1M).
Abbreviations:

Abbreviations:

DSBH,

double stranded β-helix;

FIH,

factor inhibiting HIF;

HIF,

hypoxia-inducible factor;

2OG,

2-oxoglutarate;

PHD,

prolyl hydroxylase;

SAD,

single-wavelength anomalous diffraction.