Ribosome recycling step in yeast cytoplasmic protein synthesis is catalyzed by eEF3 and ATP.
After each round of protein biosynthesis, the posttermination complex (PoTC) consisting of a ribosome, mRNA, and tRNA must be disassembled into its components for a new round of translation. Here, we show that a Saccharomyces cerevisiae model PoTC was disassembled by ATP and eukaryotic elongation factor 3 (eEF3). GTP or ITP functioned with less efficiency and adenosine 5gamma'-(beta,gamma-imido)triphosphate did not function at all. The k(cat) of eEF3 was 1.12 min(-1), which is comparable to that of the in vitro initiation step. The disassembly reaction was inhibited by aminoglycosides and cycloheximide. The subunits formed from the yeast model PoTC remained separated under ionic conditions close to those existing in vivo, suggesting that they are ready to enter the initiation process. Based on our experimental techniques used in this paper, the release of mRNA and tRNA and ribosome dissociation took place simultaneously. No 40S*mRNA complex was observed, indicating that eEF3 action promotes ribosome recycling, not reinitiation.
Department of Biochemistry and Molecular Biology, Thomas Jefferson University, 1020 Locust Street, Philadelphia, PA 19107, USA.
Shinya
S
Kurata
Klaus H
KH
Nielsen
Sarah F
SF
Mitchell
Jon R
JR
Lorsch
Akira
A
Kaji
Hideko
H
Kaji
D000255:Adenosine Triphosphate / Q000378:metabolism*; D003593:Cytoplasm / Q000378:metabolism; D007700:Kinetics; D008954:Models, Biological; D008958:Models, Molecular; D010445:Peptide Elongation Factors / Q000737:chemistry / Q000378:metabolism*; D012331:RNA, Fungal / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D012343:RNA, Transfer / Q000378:metabolism; D012270:Ribosomes / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000096:biosynthesis* / Q000737:chemistry / Q000378:metabolism
D010445:Peptide Elongation Factors; D012331:RNA, Fungal; D012333:RNA, Messenger; D029701:Saccharomyces cerevisiae Proteins; D000255:Adenosine Triphosphate; D012343:RNA, Transfer