Antibodies to Eukaryotic Initiation Factors (eIF)

Introduction

Initiation factors (IF) are one of a group of regulatory proteins that play an important role in the regulation of translation as they promote the proper association of the ribosome and the mRNA at the start of protein synthesis.

For instance, the eIF2 protein, which is composed of three subunits (α, β and γ), is one of the key molecules in the initiation of translation. In mammalian cells, stress induces eIF2α to be phosphorylated at serine 52 by at least two kinases. Phosphorylation of eIF2α blocks the GDP-GTP exchange activity of eIF2β, leading to inhibition of protein synthesis, growth suppression, and apoptosis induction.

Eukaryotic Initiation Factor 2B (eIF2B) acts as a guanine nucleotide exchange factor. The activity of eIF2B is regulated by a wide array of conditions. Stress inhibits its action, while stimulation of protein synthesis results in its activation. eIF2Bε, an 82 kDa subunit of eIF2B, is phosphorylated at serine 539 by glycogen synthase kinase-3 (GSK-3), which leads to inhibition of its guanine nucleotide exchange activity.

Eukaryotic initiation factor 4G (eIF4G) is a member of the eIF4 class of translational initiation factors involved in recognition of the mRNA cap, ATP-dependent unwinding of 5’-terminal secondary structure, and recruitment of mRNA to the ribosome. EIF4EL3 recognizes and binds the 7-methylguanosine-containing mRNA cap during an early step in the initiation of protein synthesis and facilitates ribosome binding by inducing the unwinding of the mRNAs secondary structures.

Eukaryotic translation initiation factor 5A (eIF5A) is an essential protein tightly linked to cellular polyamine homeostasis. EIF5A2, a novel phylogenetically conserved gene for eIF5A and localized in an area of the genome is often noted for chromosomal instability in cancers.

Antibody Panel to eIF

Acris Antibodies offers a range of polyclonal antibodies to various eukaryotic Initiation Factors; some of them are specific for the respective phosphorylation sites.

All antibodies are suitable for Western blot applications.

Fig. 1: Peptide competition experiment using antibody Cat.No. BP7052 (anti-eIF-2A): Extracts prepared from 3T3-L1 adipocytes treated with LIF were resolved by SDS-PAGE on a 10% polyacrylamide gel and transferred to PVDF. Membranes were blocked with a 5% nonfat dried milk-TBST buffer overnight at 4oC, then were incubated with eIF2a [pSer52] antibody for two hours at room temperature in a 3% non-fat dried milk-TBST buffer, following prior incubation with: no peptide (1), the non-phosphopeptide corresponding to the immunogen (2), a generic phosphoserine-containing peptide (3), or, the phosphopeptide immunogen (4). After washing, membranes were incubated with goat F(ab’)2 anti-rabbit IgG HRP conjugate and bands were detected using the’ Pierce SuperSignal‘ method. The data show that only the peptide corresponding to eIF2a [pSer52] blocks the antibody signal, thereby demonstrating the specificity of the antibody.

Fig.2 : Western blot analysis of 4EBP1, using antibody Cat.No. NB200-157. Samples: Whole cell lysate (100 µg) from RINm5F insulinoma exposed in high or low concentration of leucine.

Table of Antibody Panel to eIF

eukaryotic Initiation Factors (eIF) antibody.

Literature

Zoll WL, Horton LE, Komar AA, Hensold JO, Merrick WC Characterization of mammalian eIF2A and identification of the yeast homolog. J. Biol Chem. 2002 Oct 4; 277(40): 37079-87.

Telemann AA, Chen YW, Cohen SM. 4E-BP functions as a metabolic brake used under stress conditions but not during normal growth. Genes Dev. 2005 Aug 15; 19(16): 1844-8.

Clement PM, Henderson CA, Jenkins ZA, Smit-McBride Z, Wolff EC, Hershey JW, Park MH, Johansson HE. Identification and characterization of eukaryotic initiation factor 5A-2. Eur J Biochem. 2003 Nov; 270(21). 4254-63.

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