Reconstitution of yeast translation initiation.
To facilitate the mechanistic dissection of eukaryotic translation initiation we have reconstituted the steps of this process using purified Saccharomyces cerevisiae components. This system provides a bridge between biochemical studies in vitro and powerful yeast genetic techniques, and complements existing reconstituted mammalian translation systems (Benne and Hershey, 1978; Pestova and Hellen, 2000; Pestova et al., 1998; Trachsel et al., 1977). The following describes methods for synthesizing and purifying the components of the yeast initiation system and assays useful for its characterization.
Acker
Michael G
MG
Department of Biophysics and Biophysical Chemistry, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA.
Kolitz
Sarah E
SE
Mitchell
Sarah F
SF
Nanda
Jagpreet S
JS
Lorsch
Jon R
JR
D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015977:Eukaryotic Initiation Factor-1 / Q000302:isolation & purification / Q000378:metabolism; D015852:Eukaryotic Initiation Factor-2 / Q000302:isolation & purification / Q000378:metabolism; D008715:Methionine / Q000378:metabolism; D008718:Methionine-tRNA Ligase / Q000302:isolation & purification / Q000378:metabolism; D014176:Protein Biosynthesis; D020033:Protein Isoforms / Q000302:isolation & purification / Q000378:metabolism; D012331:RNA, Fungal / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D012335:RNA, Ribosomal / Q000302:isolation & purification / Q000378:metabolism; D012358:RNA, Transfer, Met / Q000378:metabolism; D054683:Ribosome Subunits, Large, Eukaryotic / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D054682:Ribosome Subunits, Small, Eukaryotic / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism
D015977:Eukaryotic Initiation Factor-1; D015852:Eukaryotic Initiation Factor-2; D020033:Protein Isoforms; D012331:RNA, Fungal; D012335:RNA, Ribosomal; D012358:RNA, Transfer, Met; D029701:Saccharomyces cerevisiae Proteins; D008715:Methionine; D008718:Methionine-tRNA Ligase