Regulation of the initiation of mrna translation by phosphorylation of eif2: Role of the α-subunit of eif-2b

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Abstract

Modulation of the activity of eukaryotic initiation factor (eIF)2B is an important means of regulating the initiation of mRNA translation. The best characterized form of regulation of eIF2B activity involves phosphorylation of the a-subunit of its substrata, eIF2. Kinetic analysis of the guanine nucleotide exchange (GEF) reaction mediated by eIF2B suggests that phosphorylation of eIF2 converts eIF2 from a substrata into a competitive inhibitor. Genetic studies in yeast suggest that the a-subunit of eIF2B plays an important role in this type of regulation, although its mode of action has not been defined biochemically. In the present study, we have coexpressed all five subunits of rat eIF2B in Sf21 cells using the baculovirus system and have purified the expressed protein to 98% homogeneity. In addition, we have expressed and purified a 4-subunit elF2B complex lacking the a-subunit. Both forms ofeIF2B exhibit similar rates of guanine nucleotide exchange using unphosphorylated eIF2 as substrata. The 5-subunit form of eIF2B is inhibited by preincubation with phosphorylated eIF2 and exhibits little GEF activity when phosphorylated elF2 is used as substrate. In contrast, eIF2B lacking the a-subunit is much less sensitive to inhibition by phosphorylated eIF2 and is able to exchange guanine nucleotide using phosphorylated eIF2 as substrate at a faster rate compared to the 5-subunit form of the protein. The results demonstrate for the first time that the a-subunit of eIF2B plays an important role in the regulation of the GEF activity of eIF2B in response to phosphorylation of eIF2.

Original languageEnglish (US)
Pages (from-to)A1403
JournalFASEB Journal
Volume11
Issue number9
StatePublished - Dec 1 1997

All Science Journal Classification (ASJC) codes

  • Biotechnology
  • Biochemistry
  • Molecular Biology
  • Genetics

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