title: Homeostasis in mRNA initiation: wheat germ poly(A)-binding protein lowers the activation energy barrier to initiation complex formation.
abstract: Previous kinetic binding studies of wheat germ protein synthesis eukaryotic initiation factor iso4F (eIFiso4F) and its subunit, eIF4E, with m(7)GTP and mRNA analogues indicated that binding occurred by a two-step process with the first step being too fast to measure by stopped-flow techniques (). Further equilibrium studies showed that poly(A)-binding protein (PABP) enhanced the cap binding of eIFiso4F about 40-fold. The kinetic effects of PABP on cap binding and the temperature dependence of this reaction were measured and compared. Fluorescence stopped-flow studies of the PABP.eIFiso4F protein complex with cap show a concentration-independent conformational change. PABP did not significantly increase the rate of the conformational change, and because the initial second-order binding is essentially diffusion-controlled, the enhancement of cap affinity must reside in the dissociation rate. The dissociation rate was more than 5-fold slower in the presence of PABP. The temperature dependence of the cap binding reaction was markedly reduced in the presence of PABP. The reduced energy barrier for formation of a cap.eIFiso4F complex suggests a more stable platform for further initiation complex formation and a possible means of adapting to varying temperature conditions.
authors.affiliation: Department of Chemistry, Hunter College and the Graduate Center of the City University of New York, New York, New York 10021, USA.
authors.forename: Y
authors.initials: Y
authors.lastname: Luo
authors.forename: D J
authors.initials: DJ
authors.lastname: Goss
mesh_terms: D039561:Eukaryotic Initiation Factor-4E; D039562:Eukaryotic Initiation Factor-4F; D006160:Guanosine Triphosphate / Q000378:metabolism; D007700:Kinetics; D008956:Models, Chemical; D010448:Peptide Initiation Factors / Q000378:metabolism; D039102:Poly(A)-Binding Proteins; D011485:Protein Binding; D012333:RNA, Messenger / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000737:chemistry* / Q000378:metabolism*; D013050:Spectrometry, Fluorescence; D013696:Temperature; D013997:Time Factors; D014908:Triticum / Q000378:metabolism*
chemical_list: D039561:Eukaryotic Initiation Factor-4E; D039562:Eukaryotic Initiation Factor-4F; D010448:Peptide Initiation Factors; D039102:Poly(A)-Binding Proteins; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D006160:Guanosine Triphosphate