TY - JOUR
T1 - Distinct functions of eukaryotic translation initiation factors eIF1A and eIF3 in the formation of the 40 S ribosomal preinitiation complex
AU - Chaudhuri, Jayanta
AU - Chowdhury, Dipanjan
AU - Maitra, Umadas
PY - 1999/6/18
Y1 - 1999/6/18
N2 - We have used an in vitro translation initiation assay to investigate the requirements for the efficient transfer of Met-tRNA(f) (as Met- tRNA(f)·eIF2·GTP ternary complex) to 40 S ribosomal subunits in the absence of mRNA (or an AUG codon) to form the 40 S preinitiation complex. We observed that the 17-kDa initiation factor eIF1A is necessary and sufficient to mediate nearly quantitative transfer of Met-tRNA(f) to isolated 40 S ribosomal subunits. However, the addition of 60 S ribosomal subunits to the 40 S preinitiation complex formed under these conditions disrupted the 40 S complex resulting in dissociation of Met-tRNA(f) from the 40 S subunit. When the eIF1A-dependent preinitiation reaction was carried out with 40 S ribosomal subunits that had been preincubated with eIF3, the 40 S preinitiation complex formed included bound eIF3 (40 S·IF3·Met- tRNA(f)·eIF2·GTP). In contrast to the complex lacking eIF3, this complex was not disrupted by the addition of 60 S ribosomal subunits. These results suggest that in vivo, both eIF1A and eIF3 are required to form a stable 40 S preinitiation complex, eIF1A catalyzing the transfer of Met- tRNA(f)·eIF2·GTP to 40 S subunits, and eIF3 stabilizing the resulting complex and preventing its disruption by 60 S ribosomal subunits.
AB - We have used an in vitro translation initiation assay to investigate the requirements for the efficient transfer of Met-tRNA(f) (as Met- tRNA(f)·eIF2·GTP ternary complex) to 40 S ribosomal subunits in the absence of mRNA (or an AUG codon) to form the 40 S preinitiation complex. We observed that the 17-kDa initiation factor eIF1A is necessary and sufficient to mediate nearly quantitative transfer of Met-tRNA(f) to isolated 40 S ribosomal subunits. However, the addition of 60 S ribosomal subunits to the 40 S preinitiation complex formed under these conditions disrupted the 40 S complex resulting in dissociation of Met-tRNA(f) from the 40 S subunit. When the eIF1A-dependent preinitiation reaction was carried out with 40 S ribosomal subunits that had been preincubated with eIF3, the 40 S preinitiation complex formed included bound eIF3 (40 S·IF3·Met- tRNA(f)·eIF2·GTP). In contrast to the complex lacking eIF3, this complex was not disrupted by the addition of 60 S ribosomal subunits. These results suggest that in vivo, both eIF1A and eIF3 are required to form a stable 40 S preinitiation complex, eIF1A catalyzing the transfer of Met- tRNA(f)·eIF2·GTP to 40 S subunits, and eIF3 stabilizing the resulting complex and preventing its disruption by 60 S ribosomal subunits.
UR - http://www.scopus.com/inward/record.url?scp=0033580851&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=0033580851&partnerID=8YFLogxK
U2 - 10.1074/jbc.274.25.17975
DO - 10.1074/jbc.274.25.17975
M3 - Article
C2 - 10364246
AN - SCOPUS:0033580851
SN - 0021-9258
VL - 274
SP - 17975
EP - 17980
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
IS - 25
ER -