title: eRF1aMC and Mg(2+) dependent structure switch of GTP binding to eRF3 in Euplotes octocarinatus.
abstract: Eukaryotic translation termination is governed by eRF1 and eRF3. eRF1 recognizes the stop codons and then hydrolyzes peptidyl-tRNA. eRF3, which facilitates the termination process, belongs to the GTPase superfamily. In this study, the effect of the MC domain of eRF1a (eRF1aMC) on the GTPase activity of eRF3 was analyzed using fluorescence spectra and high-performance liquid chromatography. The results indicated eRF1aMC promotes the GTPase activity of eRF3, which is similar to the role of eRF1a. Furthermore, the increased affinity of eRF3 for GTP induced by eRF1aMC was dependent on the concentration of Mg(2+). Changes in the secondary structure of eRF3C after binding GTP/GDP were detected by CD spectroscopy. The results revealed changes of conformation during formation of the eRF3C·GTP complex that were detected in the presence of eRF1a or eRF1aMC. The conformations of the eRF3C·eRF1a·GTP and eRF3C·eRF1aMC·GTP complexes were further altered upon the addition of Mg(2+). By contrast, there was no change in the conformation of GTP bound to free eRF3C or the eRF3C·eRF1aN complex. These results suggest that alterations in the conformation of GTP bound to eRF3 is dependent on eRF1a and Mg(2+), whereas the MC domain of eRF1a is responsible for the change in the conformation of GTP bound to eRF3 in Euplotes octocarinatus.
authors.affiliation: Institute of Biotechnology, Key Laboratory of Chemical Biology and Molecular Engineering of Ministry of Education, Shanxi University, Taiyuan 030006, China.
authors.forename: Li
authors.initials: L
authors.lastname: Song
authors.forename: Yu-Xin
authors.initials: YX
authors.lastname: Jia
authors.forename: Wen-Si
authors.initials: WS
authors.lastname: Zhu
authors.forename: Bao-Feng
authors.initials: BF
authors.lastname: Chai
authors.forename: Ai-Hua
authors.initials: AH
authors.lastname: Liang
mesh_terms: D000494:Allosteric Regulation; D002413:Cations, Divalent / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D002942:Circular Dichroism; D016812:Euplotes / Q000737:chemistry* / Q000378:metabolism*; D006160:Guanosine Triphosphate / Q000378:metabolism*; D008274:Magnesium / Q000378:metabolism*; D010454:Peptide Termination Factors / Q000737:chemistry* / Q000378:metabolism*; D011485:Protein Binding; D011487:Protein Conformation; D013050:Spectrometry, Fluorescence
chemical_list: D002413:Cations, Divalent; D010454:Peptide Termination Factors; D006160:Guanosine Triphosphate; D008274:Magnesium