Structural requirements of the mRNA for intracistronic translation initiation of the enterobacterial infB gene.
BACKGROUND
The gene infB encodes the prokaryotic translation initiation factor IF2, a central macromolecular component in the formation of the ribosomal 70S initiation complex. In Escherichia coli, infB encodes three forms of IF2: IF2alpha, IF2beta and IF2gamma. The expression of IF2beta and IF2gamma is a tandem translation from intact infB mRNA and not merely a translation of post-transcriptionally truncated mRNA. The molecular mechanism responsible for the ribosomal recognition of the two intracistronic translation initiation sites in E. coli infB is not well characterized.


RESULTS
We found three different forms of IF2 in Enterobacter cloacae, Klebsiella oxytoca, Salmonella enterica, Salmonella typhimurium, and two different forms in Proteus vulgaris. We identified the intracistronic translation initiation sites of the mRNA by isolation and N-terminal sequencing of the shorter isoforms of IF2 in S. enterica and S. typhimurium. A further search in the readily available public sequence databases revealed that infB from Yersinia pestis also contains an intracistronic in-frame initiation site used for the translation of IF2beta. The base composition in a part of the 5' end of the DNA coding strand of the enterobacterial infB gene shows a strong preference for adenine (A) over thymine (T) with a maximum ratio of A-to-T around the intracistronic initiation sites. We demonstrate that the mRNA has an open structure around the ribosomal binding region.


CONCLUSIONS
Efficient intracistronic translation initiation of the infB gene is suggested to require an mRNA with this special base composition that results in an open, single-stranded structure at the ribosomal binding region.
Laursen
Brian Søgaard
BS
Laboratory of BioDesign, Department of Molecular and Structural Biology, Aarhus University, Gustav Wieds Vej 10C, DK-8000 Aarhus C, Denmark.
de A  Steffensen
Søren A
SA
Hedegaard
Jakob
J
Moreno
Juan Manuel Palacios
JM
Mortensen
Kim Kusk
KK
Sperling-Petersen
Hans Uffe
HU
D001426:Bacterial Proteins / Q000235:genetics; D001483:Base Sequence; D004754:Enterobacter / Q000235:genetics; D004755:Enterobacteriaceae / Q000235:genetics* / Q000378:metabolism; D005798:Genes, Bacterial; D041121:Klebsiella oxytoca / Q000235:genetics; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D010442:Peptide Chain Initiation, Translational; D039881:Prokaryotic Initiation Factor-2 / Q000235:genetics*; D014176:Protein Biosynthesis; D020033:Protein Isoforms / Q000235:genetics; D011514:Proteus vulgaris / Q000235:genetics; D012329:RNA, Bacterial / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D019779:Salmonella enterica / Q000235:genetics; D016415:Sequence Alignment
D001426:Bacterial Proteins; D039881:Prokaryotic Initiation Factor-2; D020033:Protein Isoforms; D012329:RNA, Bacterial; D012333:RNA, Messenger