title: Structural and mechanistic studies on chloroplast translational initiation factor 3 from Euglena gracilis.
abstract: Chloroplast translational initiation factor 3 (IF3chl) from Euglena gracilis contains a central region (homology domain) that is homologous to prokaryotic IF3. The homology domain is preceded by a long NH2-terminal extension (head), and followed by a 64 amino acid COOH-terminal extension (tail). Sequences in these extensions reduce the activity of the homology domain. To gain insight into these effects, a possible three-dimensional structure for the homology region of IF3chl has been modeled using the X-ray coordinates from the N- and C-domains of Bacillus stearothermophilus IF3. In B. stearothermophilus IF3, these two compact domains are thought to fold independently and are separated by a helical lysine-rich linker. The modeled structure suggests that IF3chl has a similar overall fold although some subtle differences are predicted to occur. Both the head and tail regions of IF3chl are oriented toward the linker region in the homology domain where they may potentially interfere with its function. Circular dichroism spectropolarimetry (CD) indicates that the lysine-rich linker region in IF3chl is not in a helical conformation and is probably a random coil. CD analysis indicates that a portion of the tail region of IF3chl is helical and that the tail has a direct interaction with the linker region in the homology domain. Site-directed mutagenesis of the linker indicates that two conserved lysine residues are important for the function of IF3chl and play a role in the binding of IF3chl to the 30S ribosomal subunit. Mutation of these residues affects the interaction of the homology domain with the tail.
authors.affiliation: Department of Chemistry, University of North Carolina, Chapel Hill 27599-3290, USA.
authors.forename: N J
authors.initials: NJ
authors.lastname: Yu
authors.forename: L L
authors.initials: LL
authors.lastname: Spremulli
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002736:Chloroplasts; D002942:Circular Dichroism; D003198:Computer Simulation; D016384:Consensus Sequence; D005056:Euglena gracilis; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016296:Mutagenesis; D010448:Peptide Initiation Factors / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D039665:Prokaryotic Initiation Factor-3; D011485:Protein Binding; D011489:Protein Denaturation; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D012270:Ribosomes / Q000378:metabolism
chemical_list: D010448:Peptide Initiation Factors; D039665:Prokaryotic Initiation Factor-3; D011994:Recombinant Proteins