Search

The initiation of translation in eukaryotes generally involves the recognition of a 'cap' structure at the 5' end of the mRNA. However, for some viral and cellular mRNAs, a cap-independent mechanism occurs through an mRNA structure known as the internal ribosome entry site (IRES). Here, I postulate that the first eukaryotic mRNAs were translated in a cap-independent, IRES-driven manner that was then superseded in evolution by the cap-dependent mechanism, rather than vice versa. This hypothesis is supported by the following observations: (i) IRES-dependent, but not cap-dependent, translation can take place in the absence of not only a cap, but also many initiation factors; (ii) eukaryotic initiation factor 4E (elF4E) and elF4G, molecules absolutely required for cap-dependent translation, are among the most recently evolved translation factors; and (iii) functional similarities suggest the evolution of IRESs from spliceosomal introns. Thus, the contemporary cellular IRESs might be relics of the past.

A study was conducted to explore the structural properties, mechanistic details, and physiological significance of tmRNA and its principal protein partners. The evidence indicates that in addition to its quality-control function the tmRNA system might also play a key regulatory role in certain physiological pathways.

A comparative sequence-structure analysis of hfq genes in bacterial genomes is employed to identify conserved residues that may be involved in binding RNA. A covariance of residue properties at neighboring positions 12 and 39 and conserved surfaces residues with high propensities at binding sites of RNA- binding proteins suggested several sites for Hfq-RNA interactions.

A cell-free translation mixture and a 27 MHz quartz-crystal microbalance (QCM) was used to develop a system that allowed real-time monitoring of protein synthesis and understand the reactions of protein translation. The developed approach allowed assessing the effects of different antibiotics and mRNA sequences on the different steps of protein translation and the rate-limiting steps in protein synthesis.

The small size and domain mobility has made the PBCV-1 mRNA capping enzyme an ideally suited model system for exploring domain mobility in context of substrate binding. The results have shown that binding efficiency is a function of conformation but that isomerization between efficient and inefficient binding conformations have not affected the substrate association rate.

Hairpin loops from c-src and GABAA [gamma]2 pre-mRNAs and from the 3' terminus of hepatitis C virus (HCV) were compared to a single-stranded polypyrimidine tract from GABAA [gamma]2 pre-mRNA to understand the modular binding function of structured or unstructured polypyrimidine tract binding protein 1 (PTB1) RNA. The structured or single-stranded binding of short and long polypyrimidine tracts provided insights into the versatile role of PTB as an adaptor protein that facilitates formation of RNA-protein regulatory complexes.

A study was conducted to probe the crystal structures and CstF-50 binding characteristics of BARD1 C-terminal ankyrin and tandem BRCT repeat domains which modulate mRNA processing and RNAP II stability in response to DNA damage. The results revealed that BARD1 structure and plasticity imparted by the ANK-BRCT liner facilitate the BARD1 C-terminus to act as a hub with multiple binding sites to integrate diverse DNA damage signals directly to RNA polymerase.

The kinetic and thermodynamic characterization of the reaction catalyzed by an RNA guanylyltransferase capping enzyme involved in the synthesis of the 5' end of eukaryotic mRNAs, from Paramecium bursaria Chorella virus 1 is described. The findings provide insight into the highest energy required by the RNA substrates to promote the catalytic activities of the enzyme.

A comparative analysis of bacterial mRNA sequences of Bacillus stearothermophilus were carried out to define the features recognized by binding site of an unusual ribosomal protein complex, L10[(L12).sub.4]. The mRNA and ribosome targets were found to use similar RNA features, held in different structural contexts, to recognize L10[(L12).sub.4], while the ribosome is found to ensure the saturation of its L10[(L12).sub.4] binding site by means of an additional protein-protein interaction.

A combination of the hydrogen-deuterium exchange method along with electrospray mass spectrometry is applied to monitor structural rearrangement of eukaryotic initiation factor 4E (eIF4E) upon binding of the mRNA 5' cap analogues. The results suggest that the additional double methylation at the [N.sup.2]-amino group of the cap, which causes sterical effects upon binding to mammalian eIF4E, influences the overall solution dynamics of the protein.