The objective of this proposal is to elucidate the molecular mechanism in the regulation by heme of the eIF-2-alpha kinase (HRI). The specific claims include (1) determination of the molecular mechanisms by which heme regulates the autokinase and eIF-2 alpha kinase activities of HRI; (2) cloning and sequencing of HRI cDNA; comparison of sequences of HRI to those of dsRNA- dependent eIF-2 alpha kinase, and other protein kinases and heme- binding proteins; (3) determination of the genomic structure and sequences of HRI; comparison of the non-coding sequences of HRI to those of genes whose expressions are regulated by heme. HRI will be purified and assays of protein phosphorylation and protein synthesis carried out. The techniques to be used include gel electrophoresis, amino acid and nucleotide sequencing, nucleic acid hybridization and recombinant DNA. The proposed studies will advance our knowledge of the control of protein synthesis, a process that is central to many aspects of the activities of a living organism, such as the regulation of cell growth, differentiation, antiviral response and hormonal response.
