The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Use a concentration of 1 µg/ml. Predicted molecular weight: 71 kDa. Good results were obtained when blocked with 5% non-fat dry milk in 0.05% PBS-T.
Use at an assay dependent concentration.
FunctionInhibits protein synthesis at the translation initiation level, in response to various stress conditions, including oxidative stress, heme deficiency, osmotic shock and heat shock. Exerts its function through the phosphorylation of EIF2S1 at 'Ser-48' and 'Ser-51', thus preventing its recycling. Binds hemin forming a 1:1 complex through a cysteine thiolate and histidine nitrogenous coordination. This binding occurs with moderate affinity, allowing it to sense the heme concentration within the cell. Thanks to this unique heme-sensing capacity, plays a crucial role to shut off protein synthesis during acute heme-deficient conditions. In red blood cells (RBCs), controls hemoglobin synthesis ensuring a coordinated regulation of the synthesis of its heme and globin moieties. Thus plays an essential protective role for RBC survival in anemias of iron deficiency. Similarly, in hepatocytes, involved in heme-mediated translational control of CYP2B and CYP3A and possibly other hepatic P450 cytochromes. May also contain ER stress during acute heme-deficient conditions.
Tissue specificityExpressed predominantly in erythroid cells. At much lower levels, expressed in hepatocytes (at protein level).
Sequence similaritiesBelongs to the protein kinase superfamily. Ser/Thr protein kinase family. GCN2 subfamily. Contains 2 HRM (heme regulatory motif) repeats. Contains 1 protein kinase domain.
Post-translational modificationsActivated by autophosphorylation; phosphorylated predominantly on serine and threonine residues, but also on tyrosine residues. Autophosphorylation at Thr-488 is required for kinase activation. The active autophosphorylated form apparently is largely refractory to cellular heme fluctuations.