Structural requirements for porphyrin inhibition of the hemin-controlled protein kinase and maintenance of protein synthesis in reticulocytes.
The role of hemin in the maintenance of protein synthesis in reticulocyte lysates was examined by comparing the effects of various porphyrins and metalloporphyrins on the protein kinase activity of the hemin-controlled repressor and on protein synthesis. The porphyrin requirements for maintenance of protein synthesis were relatively specific. Iron and cobalt metalloporphyrins sustained protein synthesis whereas other metalloporphyrins, metal-deficient porphyrins, and non-porphyrin precursor and degradation products of protoporphyrin IX were ineffective. These same compounds were examined for their effectiveness in inhibiting the protein kinase activity of the hemin-controlled repressor with initiation factor 2 (eIF-2). Most of the metalloporphyrins and porphyrins tested were inhibitory. The presence of the iron atom in the porphyrin was not essential for inhibition, but the maintenance of the integrity of the porphyrin ring was imperative. The porphyrins which inhibited the hemin-regulated protein kinase contained vinyl groups or ethyl groups, or were protonated in the 2- and 4-positions of the porphyrin ring, whereas those with bulky or acidic groups in these positions were ineffective. Precursor and degradation products of protoporphyrin IX and synthetic porphyrins modified at other positions had no effect on the enzyme. Both hemin and protoporphyrin IX inhibited phosphorylation of eIF-2 exogenously added to a reticulocyte lysate; however, hemin sustained protein synthesis in the lysate, whereas protoporphyrin IX did not. These results suggest that regulation of the protein kinase phosphorylating the alpha subunit of eIF-2 is not the only point at which hemin modulates protein synthesis in reticulocytes and reticulocyte lysates, since a correlation between inhibition of protein synthesis, inhibition of protein kinase activity, and phosphorylation of eIF-2 is not observed with all porphyrins.
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D000818:Animals; D003035:Cobalt / Q000378:metabolism; D003306:Coproporphyrins / Q000378:metabolism; D015852:Eukaryotic Initiation Factor-2; D006418:Heme / Q000031:analogs & derivatives*; D006427:Hemin / Q000378:metabolism*; D008274:Magnesium / Q000378:metabolism; D010448:Peptide Initiation Factors / Q000378:metabolism; D010766:Phosphorylation; D011166:Porphyrins / Q000494:pharmacology*; D014176:Protein Biosynthesis; D047428:Protein Kinase Inhibitors; D011506:Proteins / Q000378:metabolism; D011524:Protoporphyrins / Q000494:pharmacology; D011817:Rabbits; D012156:Reticulocytes / Q000378:metabolism*; D015032:Zinc / Q000378:metabolism
D003306:Coproporphyrins; D015852:Eukaryotic Initiation Factor-2; D010448:Peptide Initiation Factors; D011166:Porphyrins; D047428:Protein Kinase Inhibitors; D011506:Proteins; D011524:Protoporphyrins; C026971:coproporphyrin III; D003035:Cobalt; D006418:Heme; D006427:Hemin; C028025:protoporphyrin IX; D008274:Magnesium; D015032:Zinc