Phosphorylation of Serine 51 in Initiation Factor 2 (eIF2) Promo
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- 作者：Akulapalli Sudhakar ; Aruna Ramachandran ; Sudip Ghosh ; Seyed E. Hasnain ; Randal J. Kaufman ; and Kolluru V. A. Ramaiah
- 刊名：Biochemistry
- 出版年：2000
- 出版时间：October 24, 2000
- 年：2000
- 卷：39
- 期：42
- 页码：12929 - 12938
- 全文大小：213K
- 年卷期：v.39,no.42(October 24, 2000)
- ISSN：1520-4995
文摘
Phosphorylation of serine 51 residue on the 
-subunit of eukaryotic initiation factor 2 (eIF2)inhibits the guanine nucleotide exchange (GNE) activity of eIF2B, presumably, by forming a tight complexwith eIF2B. Inhibition of the GNE activity of eIF2B leads to impairment in eIF2 recycling and proteinsynthesis. We have partially purified the wild-type (wt) and mutants of eIF2 in which the serine 51residue was replaced with alanine (51A mutant) or aspartic acid (51D mutant) in the baculovirus system.Analysis of these mutants has provided novel insight into the role of 51 serine in the interaction betweeneIF2 and eIF2B. Neither mutant was phosphorylated in vitro. Both mutants decreased eIF2 phosphorylationoccurring in hemin and poly(IC)-treated reticulocyte lysates due to the activation of double-stranded RNA-dependent protein kinase (PKR). However, addition of 51D, but not 51A mutant eIF2 protein promotedinhibition of the GNE activity of eIF2B in hemin-supplemented rabbit reticulocyte lysates in which relativelylittle or no endogenous eIF2 phosphorylation occurred. The 51D mutant enhanced the inhibition in GNEactivity of eIF2B that occurred in hemin and poly(IC)-treated reticulocyte lysates where PKR is active.Our results show that the increased interaction between eIF2 and eIF2B protein, occurring in reticulocytelysates due to increased eIF2 phosphorylation, is decreased significantly by the addition of mutant 51Aprotein but not 51D. Consistent with the idea that mutant 51D protein behaves like a phosphorylatedeIF2, addition of this partially purified recombinant subunit, but not 51A or wt eIF2, increases theinteraction between eIF2 and 2B proteins in actively translating hemin-supplemented lysates. These findingssupport the idea that phosphorylation of the serine 51 residue in eIF2 promotes complex formationbetween eIF2(P) and eIF2B and thereby inhibits the GNE activity of eIF2B.
-subunit of eukaryotic initiation factor 2 (eIF2)inhibits the guanine nucleotide exchange (GNE) activity of eIF2B, presumably, by forming a tight complexwith eIF2B. Inhibition of the GNE activity of eIF2B leads to impairment in eIF2 recycling and proteinsynthesis. We have partially purified the wild-type (wt) and mutants of eIF2 in which the serine 51residue was replaced with alanine (51A mutant) or aspartic acid (51D mutant) in the baculovirus system.Analysis of these mutants has provided novel insight into the role of 51 serine in the interaction betweeneIF2 and eIF2B. Neither mutant was phosphorylated in vitro. Both mutants decreased eIF2 phosphorylationoccurring in hemin and poly(IC)-treated reticulocyte lysates due to the activation of double-stranded RNA-dependent protein kinase (PKR). However, addition of 51D, but not 51A mutant eIF2 protein promotedinhibition of the GNE activity of eIF2B in hemin-supplemented rabbit reticulocyte lysates in which relativelylittle or no endogenous eIF2 phosphorylation occurred. The 51D mutant enhanced the inhibition in GNEactivity of eIF2B that occurred in hemin and poly(IC)-treated reticulocyte lysates where PKR is active.Our results show that the increased interaction between eIF2 and eIF2B protein, occurring in reticulocytelysates due to increased eIF2 phosphorylation, is decreased significantly by the addition of mutant 51Aprotein but not 51D. Consistent with the idea that mutant 51D protein behaves like a phosphorylatedeIF2, addition of this partially purified recombinant subunit, but not 51A or wt eIF2, increases theinteraction between eIF2 and 2B proteins in actively translating hemin-supplemented lysates. These findingssupport the idea that phosphorylation of the serine 51 residue in eIF2 promotes complex formationbetween eIF2(P) and eIF2B and thereby inhibits the GNE activity of eIF2B.