Anti-HIF-1-alpha (hydroxy P402) antibody (ab72775)
- Product nameAnti-HIF-1-alpha (hydroxy P402) antibodySee all HIF-1-alpha primary antibodies ...
- DescriptionRabbit polyclonal to HIF-1-alpha (hydroxy P402)
- SpecificityThis antibody reacts specifically with HIF-1-alpha (Hydroxy P402). No crossreactivity was shown with non modified peptide.
- Tested applicationsWB, ICC/IF more details
- Species reactivityReacts with: Human
Predicted to work with: a wide range of other species
Synthetic peptide corresponding to Human HIF-1-alpha.
- Positive control
- IF/ICC: HeLa cell line.
- Storage instructionsStore at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C. Avoid freeze / thaw cycle.
- Storage bufferConstituent: Whole serum
- PurityWhole antiserum
- Clonality Polyclonal
Our Abpromise guarantee covers the use of ab72775 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|WB||WB: 1/200 - 1/1000. Predicted molecular weight: 93 kDa.|
- FunctionFunctions as a master transcriptional regulator of the adaptive response to hypoxia. Under hypoxic conditions activates the transcription of over 40 genes, including, erythropoietin, glucose transporters, glycolytic enzymes, vascular endothelial growth factor, and other genes whose protein products increase oxygen delivery or facilitate metabolic adaptation to hypoxia. Plays an essential role in embryonic vascularization, tumor angiogenesis and pathophysiology of ischemic disease. Binds to core DNA sequence 5'-[AG]CGTG-3' within the hypoxia response element (HRE) of target gene promoters. Activation requires recruitment of transcriptional coactivators such as CREBPB and EP300. Activity is enhanced by interaction with both, NCOA1 or NCOA2. Interaction with redox regulatory protein APEX seems to activate CTAD and potentiates activation by NCOA1 and CREBBP.
- Tissue specificityExpressed in most tissues with highest levels in kidney and heart. Overexpressed in the majority of common human cancers and their metastases, due to the presence of intratumoral hypoxia and as a result of mutations in genes encoding oncoproteins and tumor suppressors.
- Sequence similaritiesContains 1 basic helix-loop-helix (bHLH) domain.
Contains 1 PAC (PAS-associated C-terminal) domain.
Contains 2 PAS (PER-ARNT-SIM) domains.
- DomainContains two independent C-terminal transactivation domains, NTAD and CTAD, which function synergistically. Their transcriptional activity is repressed by an intervening inhibitory domain (ID).
modificationsIn normoxia, is hydroxylated on Pro-402 and Pro-564 in the oxygen-dependent degradation domain (ODD) by EGLN1/PHD1 and EGLN2/PHD2. EGLN3/PHD3 has also been shown to hydroxylate Pro-564. The hydroxylated prolines promote interaction with VHL, initiating rapid ubiquitination and subsequent proteasomal degradation. Deubiquitinated by USP20. Under hypoxia, proline hydroxylation is impaired and ubiquitination is attenuated, resulting in stabilization.
In normoxia, is hydroxylated on Asn-803 by HIF1AN, thus abrogating interaction with CREBBP and EP300 and preventing transcriptional activation. This hydroxylation is inhibited by the Cu/Zn-chelator, Clioquinol.
S-nitrosylation of Cys-800 may be responsible for increased recruitment of p300 coactivator necessary for transcriptional activity of HIF-1 complex.
Requires phosphorylation for DNA-binding.
Sumoylated; by SUMO1 under hypoxia. Sumoylation is enhanced through interaction with RWDD3. Desumoylation by SENP1 leads to increased HIF1A stability and transriptional activity.
Ubiquitinated; in normoxia, following hydroxylation and interaction with VHL. Lys-532 appears to be the principal site of ubiquitination. Clioquinol, the Cu/Zn-chelator, inhibits ubiquitination through preventing hydroxylation at Asn-803.
The iron and 2-oxoglutarate dependent 3-hydroxylation of asparagine is (S) stereospecific within HIF CTAD domains.
- Cellular localizationCytoplasm. Nucleus. Cytoplasmic in normoxia, nuclear translocation in response to hypoxia. Colocalizes with SUMO1 in the nucleus, under hypoxia.
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Anti-HIF-1-alpha (hydroxy P402) antibody images
ICC/IF image of ab72775 stained HeLa cells. The cells were 4% formaldehyde fixed (10 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab72775, 1/50 dilution) overnight at +4°C. The secondary antibody (green) was ab96899, DyLight® 488 goat anti-rabbit IgG (H+L) used at a 1/250 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.
Predicted band size : 93 kDa
Image from Wiley M et al., J Biol Chem. 2010 Aug 27;285(35):26852-60. Epub 2010 Jun 25. Fig 2.; doi: 10.1074/jbc.M110.147041; August 27, 2010 The Journal of Biological Chemistry, 285, 26852-26860.Representative western blot detecting HIF1 alpha (Hydroxy P402) with ab72775; a different antibody was used to detect total HIF1 alpha and an antibody against actin was used as a loading control.
Lane 1: Mock infected Human foreskin fibroblasts
Lane 2: CoCl2-treated Human foreskin fibroblasts
Lane 3: Toxoplasma-infected Human foreskin fibroblasts
Lane 4: MG132-treated Human foreskin fibroblasts
References for Anti-HIF-1-alpha (hydroxy P402) antibody (ab72775)
This product has been referenced in:
- Wiley M et al. Toxoplasma gondii activates hypoxia-inducible factor (HIF) by stabilizing the HIF-1alpha subunit via type I activin-like receptor kinase receptor signaling. J Biol Chem 285:26852-60 (2010). WB ; Human . Read more (PubMed: 20581113) »