Biophysical and Structural Characterization of the Recombinant Human eIF3L

Abstract

The eukaryotic translation initiation factor 3, subunit L (eIF3L) is one of the subunits of the eIF3 complex, an accessory protein of the Polymerase I enzyme and may have an important role in the Flavivirus replication by interaction with a viral non-structural 5 protein. Considering the importance of eIF3L in a diversity of cellular functions, we have produced the recombinant full-length eIF3L protein in Escherichia coli and performed spectroscopic and in silico analyses to gain insights into its hydrodynamic behavior and structure. Dynamic light scattering showed that eIF3L behaves as monomer when it is not interacting with other molecular partners. Circular dichroism experiments showed a typical spectrum of α-helical protein for eIF3L, which is supported by sequence-based predictions of secondary structure and the 3D in silico model. The molecular docking with the K subunit of the eIF3 complex revealed a strong interaction. It was also predicted several potential interaction sites in eIF3L, indicating that the protein is likely capable of interacting with other molecules as experimentally shown in other functional studies. Moreover, bioinformatics analyses showed approximately 8 putative phosphorylation sites and one possible N-glycosylation site, suggesting its regulation by post-translational modifications. The production of the eIF3L protein in E. coli and structural information gained in this study can be instrumental for target-based drug design and inhibitors against Flavivirus replication and to shed light on the molecular mechanisms involved in the eukaryotic translation initiation.

Toward the mechanism of eIF4F-mediated ribosomal attachment to mammalian capped mRNAs

“…eIF3l is not essential for eIF3's function (Masutani et al 2007;Morris et al 2007). eIF3l's cellular role remains uncharacterized, although, when overexpressed, it modestly represses yellow fever virus (YFV) replication (Schoggins et al 2011), binds to the YFV RNA polymerase, and specifically inhibits YFV but not general cellular translation (Morais et al 2014). We suggest that the eIF3l/cap interaction might play a regulatory role in the translation of specific mRNAs with particularly high affinity to the factor.…”

Molecular characterization and functional analysis of subunit 7 of eukaryotic initiation factor 3 from Eimeria tenella

“…These data suggest that its function might be regulated by posttranslational modification, which might regulate the activity and interactions of the subunits of the eIF3 complex during the initiation of translation. The phosphorylation of eIF3 subunits is directly involved in interactions with other subunits and other cellular components (Damoc et al, 2007;Morais et al, 2014).…”