Structural changes of eIF4E upon binding to the mRNA 5' monomethylguanosine and trimethylguanosine Cap.
Recognition of the 5' cap by the eukaryotic initiation factor 4E (eIF4E) is the rate-limiting step in the ribosome recruitment to mRNAs. The regular cap consists of 7-monomethylguanosine (MMG) linked by a 5'-5' triphosphate bridge to the first transcribed nucleoside, while some primitive eukaryotes possess a N (2), N (2),7-trimethylguanosine (TMG) cap structure as a result of trans splicing. Mammalian eIF4E is highly specific to the MMG form of the cap in terms of association constants and thermodynamic driving force. We have investigated conformational changes of eIF4E induced by interaction with two cap analogues, 7-methyl-GTP and N (2), N (2),7-trimethyl-GTP. Hydrogen-deuterium exchange and electrospray mass spectrometry were applied to probe local dynamics of murine eIF4E in the apo and cap-bound forms. The data show that the cap binding induces long-range conformational changes in the protein, not only in the cap-binding pocket but also in a distant region of the 4E-BP/eIF4G binding site. Formation of the complex with 7-methyl-GTP makes the eIF4E structure more compact, while binding of N (2), N (2),7-trimethyl-GTP leads to higher solvent accessibility of the protein backbone in comparison with the apo form. The results suggest that the additional double methylation at the N (2)-amino group of the cap causes sterical effects upon binding to mammalian eIF4E which influence the overall solution dynamics of the protein, thus precluding formation of a tight complex.
Rutkowska-Wlodarczyk
Izabela
I
Institute of Biochemistry and Biophysics, Polish Academy of Sciences, 02-106 Warsaw, Poland.
Stepinski
Janusz
J
Dadlez
Michal
M
Darzynkiewicz
Edward
E
Stolarski
Ryszard
R
Niedzwiecka
Anna
A
D000595:Amino Acid Sequence; D001665:Binding Sites; D018360:Crystallography, X-Ray; D039561:Eukaryotic Initiation Factor-4E / Q000737:chemistry* / Q000378:metabolism; D006151:Guanosine / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism; D008969:Molecular Sequence Data; D011485:Protein Binding; D012314:RNA Cap Analogs / Q000737:chemistry* / Q000378:metabolism; D039381:RNA Cap-Binding Proteins / Q000737:chemistry / Q000378:metabolism; D012333:RNA, Messenger / Q000737:chemistry* / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D013329:Structure-Activity Relationship
C116616:8-methylguanosine; D039561:Eukaryotic Initiation Factor-4E; D012314:RNA Cap Analogs; D039381:RNA Cap-Binding Proteins; D012333:RNA, Messenger; D006151:Guanosine