Functional analysis of the translation factor aIF2/5B in the thermophilic archaeon Sulfolobus solfataricus.
The protein IF2/eIF5B is one of the few translation initiation factors shared by all three primary domains of life (bacteria, archaea, eukarya). Despite its phylogenetic conservation, the factor is known to present marked functional divergences in the bacteria and the eukarya. In this work, the function in translation of the archaeal homologue (aIF2/5B) has been analysed in detail for the first time using a variety of in vitro assays. The results revealed that the protein is a ribosome-dependent GTPase which strongly stimulates the binding of initiator tRNA to the ribosomes even in the absence of other factors. In agreement with this finding, aIF2/5B enhances the translation of both leadered and leaderless mRNAs when expressed in a cell-free protein-synthesizing system. Moreover, the degree of functional conservation of the IF2-like factors in the archaeal and bacterial lineages was investigated by analysing the behaviour of 'chimeric' proteins produced by swapping domains between the Sulfolobus solfataricus aIF2/5B factor and the IF2 protein of the thermophilic bacterium Bacillus stearothermophilus. Beside evidencing similarities and differences between the archaeal and bacterial factors, these experiments have provided insight into the common role played by the IF2/5B proteins in all extant cells.
Maone
Enzo
E
Dpt. of Biotecnologie Cellulari ed Ematologia, Università di Roma Sapienza, Viale Regina Elena 324 Roma, Italy.
Di Stefano
Michele
M
Berardi
Alessandra
A
Benelli
Dario
D
Marzi
Stefano
S
La Teana
Anna
A
Londei
Paola
P
D020121:5' Untranslated Regions / Q000235:genetics; D001426:Bacterial Proteins / Q000737:chemistry / Q000378:metabolism; D001665:Binding Sites; D003001:Cloning, Molecular; D017124:Conserved Sequence; D020558:GTP Phosphohydrolases / Q000378:metabolism; D015870:Gene Expression; D019766:Genes, Archaeal; D006868:Hydrolysis; D010442:Peptide Chain Initiation, Translational; D010448:Peptide Initiation Factors / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D011485:Protein Binding; D014176:Protein Biosynthesis; D017433:Protein Structure, Secondary; D012358:RNA, Transfer, Met / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012270:Ribosomes / Q000378:metabolism; D048229:Sulfolobus solfataricus / Q000235:genetics / Q000378:metabolism*
D020121:5' Untranslated Regions; D001426:Bacterial Proteins; C118735:IF2 protein, archaeal; C112736:Met-tRNA(i)(Met); D010448:Peptide Initiation Factors; D012358:RNA, Transfer, Met; D011993:Recombinant Fusion Proteins; C024988:tRNA, formylmethionine-; D020558:GTP Phosphohydrolases