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Heme-regulated eIF-2alpha kinase purifies as a hemoprotein
P J Chefalo1, J Oh, M Rafie-Kolpin
1Harvard-MIT Division of Health Sciences and Technology, MIT, Cambridge, Massachusetts 02139, USA.
European Journal of Biochemistry
|January 5, 1999
Summary
Heme-regulated eIF-2alpha kinase (HRI) acts as a heme sensor in erythroid cells. It binds heme, regulating protein synthesis by controlling translational initiation.
Area of Science:
- Biochemistry
- Molecular Biology
- Cellular Biology
Background:
- Protein synthesis regulation by heme availability in reticulocytes is known.
- The precise mechanism of heme's regulation of translational initiation remains unclear.
Purpose of the Study:
- To investigate the direct heme regulation of homogeneous heme-regulated eIF-2alpha kinase (HRI).
- To elucidate the role of HRI as a heme sensor in erythroid cells.
Main Methods:
- Purification of HRI and characterization as a hemoprotein.
- Assay of HRI's autokinase and eIF-2alpha kinase activities.
- Investigation of heme inhibition kinetics and binding sites.
Main Results:
- Purified HRI is an active hemoprotein kinase inhibited by hemin (Ki = 0.5 microM).
- HRI exists as a homodimer with two distinct heme-binding sites.
- One site shows stable heme binding; the other mediates rapid HRI activity downregulation.
Conclusions:
- HRI directly binds heme, functioning as a sensor for heme availability.
- HRI coordinates balanced synthesis of globins and heme in erythroid cells.
- Heme binding to HRI is crucial for regulating translational initiation.