Visualization of the eEF2-80S ribosome transition-state complex by cryo-electron microscopy.
In an attempt to understand ribosome-induced GTP hydrolysis on eEF2, we determined a 12.6-A cryo-electron microscopy reconstruction of the eEF2-bound 80S ribosome in the presence of aluminum tetrafluoride and GDP, with aluminum tetrafluoride mimicking the gamma-phosphate during hydrolysis. This is the first visualization of a structure representing a transition-state complex on the ribosome. Tight interactions are observed between the factor's G domain and the large ribosomal subunit, as well as between domain IV and an intersubunit bridge. In contrast, some of the domains of eEF2 implicated in small subunit binding display a large degree of flexibility. Furthermore, we find support for a transition-state model conformation of the switch I region in this complex where the reoriented switch I region interacts with a conserved rRNA region of the 40S subunit formed by loops of the 18S RNA helices 8 and 14. This complex is structurally distinct from the eEF2-bound 80S ribosome complexes previously reported, and analysis of this map sheds light on the GTPase-coupled translocation mechanism.
Sengupta
Jayati
J
Wadsworth Center, New York State Department of Health, Empire State Plaza, Albany, NY 12201-0509, USA.
Nilsson
Jakob
J
Gursky
Richard
R
Kjeldgaard
Morten
M
Nissen
Poul
P
Frank
Joachim
J
D017607:Aluminum Compounds / Q000494:pharmacology; D020285:Cryoelectron Microscopy; D005459:Fluorides / Q000494:pharmacology; D005656:Fungal Proteins / Q000737:chemistry / Q000378:metabolism / Q000648:ultrastructure; D006153:Guanosine Diphosphate / Q000378:metabolism; D006160:Guanosine Triphosphate / Q000378:metabolism; D006868:Hydrolysis / Q000187:drug effects; D008958:Models, Molecular; D020652:Peptide Elongation Factor 2 / Q000737:chemistry / Q000378:metabolism* / Q000648:ultrastructure*; D017434:Protein Structure, Tertiary; D012270:Ribosomes / Q000187:drug effects / Q000378:metabolism* / Q000648:ultrastructure*
D017607:Aluminum Compounds; D005656:Fungal Proteins; D020652:Peptide Elongation Factor 2; D006153:Guanosine Diphosphate; C050992:tetrafluoroaluminate; D006160:Guanosine Triphosphate; D005459:Fluorides