Transcribed sequences of the Escherichia coli btuB gene control its expression and regulation by vitamin B12.
The Escherichia coli btuB gene product is an outer membrane protein required for the active transport of vitamin B12 and other cobalamins. Synthesis of BtuB is repressed when cells are grown in the presence of cobalamins. Mapping of the 5' end of the btuB transcript revealed that a 240-nucleotide transcribed leader precedes the coding sequence. Point mutations causing increased expression under repressing conditions were isolated by use of a btuB-lacZ gene fusion. Mutations at many sites within the leader region affected btuB-lacZ regulation, whereas some base changes upstream of the start of transcription affected the absolute level of expression but not its repressibility. Analysis of btuB-phoA gene fusions and btuB-lacZ operon and gene fusions of various lengths showed that sequences within the btuB coding region (between nucleotides +250 and +350) had to be present for proper expression and transcriptional regulation. Sequences within the leader region (up to +250) conferred regulation of translational fusions. These results indicate that btuB expression is controlled at both the transcriptional and translational levels and that different but possibly overlapping sequences in the transcribed region, including the coding region for the transport protein itself, mediate these two modes of regulation.
Lundrigan
M D
MD
Department of Microbiology, School of Medicine, University of Virginia, Charlottesville 22908.
Köster
W
W
Kadner
R J
RJ
D000595:Amino Acid Sequence; D001425:Bacterial Outer Membrane Proteins / Q000235:genetics*; D001483:Base Sequence; D004926:Escherichia coli / Q000187:drug effects / Q000235:genetics*; D029968:Escherichia coli Proteins; D015870:Gene Expression / Q000187:drug effects; D015964:Gene Expression Regulation, Bacterial / Q000187:drug effects; D026901:Membrane Transport Proteins; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009690:Nucleic Acid Conformation; D010957:Plasmids; D014176:Protein Biosynthesis; D011956:Receptors, Cell Surface / Q000235:genetics* / Q000302:isolation & purification; D018000:Receptors, Peptide; D011993:Recombinant Fusion Proteins / Q000302:isolation & purification; D015183:Restriction Mapping; D013816:Thermodynamics; D014158:Transcription, Genetic / Q000187:drug effects; D014805:Vitamin B 12 / Q000494:pharmacology*
D001425:Bacterial Outer Membrane Proteins; C044592:BtuB protein, E coli; D029968:Escherichia coli Proteins; D026901:Membrane Transport Proteins; D011956:Receptors, Cell Surface; D018000:Receptors, Peptide; D011993:Recombinant Fusion Proteins; D014805:Vitamin B 12