title: The cryo-EM structure of a translation initiation complex from Escherichia coli.
abstract: The 70S ribosome and its complement of factors required for initiation of translation in E. coli were purified separately and reassembled in vitro with GDPNP, producing a stable initiation complex (IC) stalled after 70S assembly. We have obtained a cryo-EM reconstruction of the IC showing IF2*GDPNP at the intersubunit cleft of the 70S ribosome. IF2*GDPNP contacts the 30S and 50S subunits as well as fMet-tRNA(fMet). IF2 here adopts a conformation radically different from that seen in the recent crystal structure of IF2. The C-terminal domain of IF2 binds to the single-stranded portion of fMet-tRNA(fMet), thereby forcing the tRNA into a novel orientation at the P site. The GTP binding domain of IF2 binds to the GTPase-associated center of the 50S subunit in a manner similar to EF-G and EF-Tu. Additionally, we present evidence for the localization of IF1, IF3, one C-terminal domain of L7/L12, and the N-terminal domain of IF2 in the initiation complex.
authors.affiliation: Howard Hughes Medical Institute, Health Research, Inc. at the Wadsworth Center, Albany, New York 12201, USA.
authors.forename: Gregory S
authors.initials: GS
authors.lastname: Allen
authors.forename: Andrey
authors.initials: A
authors.lastname: Zavialov
authors.forename: Richard
authors.initials: R
authors.lastname: Gursky
authors.forename: Måns
authors.initials: M
authors.lastname: Ehrenberg
authors.forename: Joachim
authors.initials: J
authors.lastname: Frank
mesh_terms: D020285:Cryoelectron Microscopy; D004926:Escherichia coli / Q000502:physiology* / Q000648:ultrastructure; D019605:Euryarchaeota / Q000378:metabolism; D039881:Prokaryotic Initiation Factor-2 / Q000378:metabolism; D014176:Protein Biosynthesis / Q000502:physiology*; D017434:Protein Structure, Tertiary; D012343:RNA, Transfer / Q000378:metabolism; D012270:Ribosomes / Q000378:metabolism* / Q000648:ultrastructure
chemical_list: D039881:Prokaryotic Initiation Factor-2; D012343:RNA, Transfer