Article ID | Journal | Published Year | Pages | File Type |
---|---|---|---|---|
10843499 | Protein Expression and Purification | 2005 | 10 Pages |
Abstract
Prolyl-4-hydroxylase domain-containing enzymes (PHDs) mediate the oxygen-dependent regulation of the heterodimeric transcription factor hypoxia-inducible factor-1 (HIF-1). Under normoxic conditions, one of the subunits of HIF-1, HIF-1α, is hydroxylated on specific proline residues to target HIF-1α for degradation by the ubiquitin-proteasome pathway. Under hypoxic conditions, the hydroxylation by the PHDs is attenuated by lack of the oxygen substrate, allowing HIF-1 to accumulate, translocate to the nucleus, and mediate HIF-mediated gene transcription. In several mammalian species including humans, three PHDs have been identified. We report here the cloning of a full-length rat cDNA that is highly homologous to the human and murine PHD-1 enzymes and encodes a protein that is 416 amino acids long. Both cDNA and protein are widely expressed in rat tissues and cell types. We demonstrate that purified and crude baculovirus-expressed rat PHD-1 exhibits HIF-1α specific prolyl hydroxylase activity with similar substrate affinities and is comparable to human PHD-1 protein.
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Authors
Ronald R. Cobb, John McClary, Warren Manzana, Silke Finster, Brent Larsen, Eric Blasko, Jennifer Pearson, Sara Biancalana, Katalin Kauser, Peter Bringmann, David R. Light, Sabine Schirm,