title: Possible steps of complete disassembly of post-termination complex by yeast eEF3 deduced from inhibition by translocation inhibitors.
abstract: Ribosomes, after one round of translation, must be recycled so that the next round of translation can occur. Complete disassembly of post-termination ribosomal complex (PoTC) in yeast for the recycling consists of three reactions: release of tRNA, release of mRNA and splitting of ribosomes, catalyzed by eukaryotic elongation factor 3 (eEF3) and ATP. Here, we show that translocation inhibitors cycloheximide and lactimidomycin inhibited all three reactions. Cycloheximide is a non-competitive inhibitor of both eEF3 and ATP. The inhibition was observed regardless of the way PoTC was prepared with either release factors or puromycin. Paromomycin not only inhibited all three reactions but also re-associated yeast ribosomal subunits. On the other hand, sordarin or fusidic acid, when applied together with eEF2/GTP, specifically inhibited ribosome splitting without blocking of tRNA/mRNA release. From these inhibitor studies, we propose that, in accordance with eEF3's known function in elongation, the release of tRNA via exit site occurs first, then mRNA is released, followed by the splitting of ribosomes during the disassembly of post-termination complexes catalyzed by eEF3 and ATP.
authors.affiliation: Department of Biochemistry and Molecular Biology, Kimmel Cancer Center, Thomas Jefferson University, Philadelphia, PA 19107, USA.
authors.forename: Shinya
authors.initials: S
authors.lastname: Kurata
authors.forename: Ben
authors.initials: B
authors.lastname: Shen
authors.forename: Jun O
authors.initials: JO
authors.lastname: Liu
authors.forename: Nono
authors.initials: N
authors.lastname: Takeuchi
authors.forename: Akira
authors.initials: A
authors.lastname: Kaji
authors.forename: Hideko
authors.initials: H
authors.lastname: Kaji
mesh_terms: D000255:Adenosine Triphosphate / Q000037:antagonists & inhibitors / Q000378:metabolism; D003513:Cycloheximide / Q000494:pharmacology; D005656:Fungal Proteins / Q000378:metabolism*; D005672:Fusidic Acid / Q000494:pharmacology; D007192:Indenes / Q000494:pharmacology; D018942:Macrolides / Q000494:pharmacology; D010303:Paromomycin / Q000494:pharmacology; D010443:Peptide Chain Termination, Translational / Q000187:drug effects*; D010445:Peptide Elongation Factors / Q000037:antagonists & inhibitors / Q000378:metabolism*; D010454:Peptide Termination Factors / Q000378:metabolism; D010881:Piperidones / Q000494:pharmacology; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology*; D012333:RNA, Messenger / Q000378:metabolism; D012343:RNA, Transfer / Q000378:metabolism; D012270:Ribosomes / Q000187:drug effects / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000187:drug effects / Q000235:genetics*
chemical_list: D005656:Fungal Proteins; D007192:Indenes; D018942:Macrolides; D010445:Peptide Elongation Factors; D010454:Peptide Termination Factors; D010881:Piperidones; D011500:Protein Synthesis Inhibitors; D012333:RNA, Messenger; C003329:sordarin; C077633:lactimidomycin; D005672:Fusidic Acid; D010303:Paromomycin; D000255:Adenosine Triphosphate; D012343:RNA, Transfer; D003513:Cycloheximide