Hypusine formation in protein by a two-step process in cell lysates.
The putative protein synthesis initiation factor eukaryotic initiation factor 4D (eIF-4D) is post-translationally modified by the polyamine spermidine, forming the rare amino acid hypusine from a lysine residue. The hypusine precursor, deoxyhypusine, was formed in crude cell lysates at pH 9.5 and converted to hypusine at pH 7.1. The modification occurred in eIF-4D, since the isoelectric points and molecular weights of the proteins modified in intact cells and lysates were indistinguishable. Only lysates from cells treated with alpha-difluoromethylornithine, to deplete endogenous polyamine pools, supported the formation of deoxyhypusine, suggesting that unmodified eIF-4D accumulated in spermidine deficient cells. Guazatine, an inhibitor of enzymes which form delta 1-pyrroline from spermidine, blocked deoxyhypusine formation in lysates by nearly 70% at 100 microM and completely at 1 mM. Other mammalian amine oxidase inhibitors had little or no effect on this reaction. Thus, deoxyhypusine formation in eIF-4D is catalyzed by a guazatine-sensitive enzyme with a basic pH optimum.
D000818:Animals; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008114:Liver Neoplasms, Experimental / Q000378:metabolism; D008239:Lysine / Q000031:analogs & derivatives* / Q000032:analysis; D009363:Neoplasm Proteins / Q000378:metabolism*; D010448:Peptide Initiation Factors / Q000378:metabolism*; D011073:Polyamines / Q000032:analysis*; D011499:Protein Processing, Post-Translational; D016601:RNA-Binding Proteins; D051381:Rats; D000097574:Eukaryotic Translation Initiation Factor 5A
D008239:Lysine; D009363:Neoplasm Proteins; D010448:Peptide Initiation Factors; D011073:Polyamines; D016601:RNA-Binding Proteins; D000097574:Eukaryotic Translation Initiation Factor 5A; C100028:hypusine